\\n\\n
Released this past November, the list is based on data collected from the Web of Science and highlights some of the world’s most influential scientific minds by naming the researchers whose publications over the previous decade have included a high number of Highly Cited Papers placing them among the top 1% most-cited.
\\n\\nWe wish to congratulate all of the researchers named and especially our authors on this amazing accomplishment! We are happy and proud to share in their success!
Note: Edited in March 2021
\\n"}]',published:!0,mainMedia:{caption:"Highly Cited",originalUrl:"/media/original/117"}},components:[{type:"htmlEditorComponent",content:'IntechOpen is proud to announce that 191 of our authors have made the Clarivate™ Highly Cited Researchers List for 2020, ranking them among the top 1% most-cited.
\n\nThroughout the years, the list has named a total of 261 IntechOpen authors as Highly Cited. Of those researchers, 69 have been featured on the list multiple times.
\n\n\n\nReleased this past November, the list is based on data collected from the Web of Science and highlights some of the world’s most influential scientific minds by naming the researchers whose publications over the previous decade have included a high number of Highly Cited Papers placing them among the top 1% most-cited.
\n\nWe wish to congratulate all of the researchers named and especially our authors on this amazing accomplishment! We are happy and proud to share in their success!
Note: Edited in March 2021
\n'}],latestNews:[{slug:"webinar-introduction-to-open-science-wednesday-18-may-1-pm-cest-20220518",title:"Webinar: Introduction to Open Science | Wednesday 18 May, 1 PM CEST"},{slug:"step-in-the-right-direction-intechopen-launches-a-portfolio-of-open-science-journals-20220414",title:"Step in the Right Direction: IntechOpen Launches a Portfolio of Open Science Journals"},{slug:"let-s-meet-at-london-book-fair-5-7-april-2022-olympia-london-20220321",title:"Let’s meet at London Book Fair, 5-7 April 2022, Olympia London"},{slug:"50-books-published-as-part-of-intechopen-and-knowledge-unlatched-ku-collaboration-20220316",title:"50 Books published as part of IntechOpen and Knowledge Unlatched (KU) Collaboration"},{slug:"intechopen-joins-the-united-nations-sustainable-development-goals-publishers-compact-20221702",title:"IntechOpen joins the United Nations Sustainable Development Goals Publishers Compact"},{slug:"intechopen-signs-exclusive-representation-agreement-with-lsr-libros-servicios-y-representaciones-s-a-de-c-v-20211123",title:"IntechOpen Signs Exclusive Representation Agreement with LSR Libros Servicios y Representaciones S.A. de C.V"},{slug:"intechopen-expands-partnership-with-research4life-20211110",title:"IntechOpen Expands Partnership with Research4Life"},{slug:"introducing-intechopen-book-series-a-new-publishing-format-for-oa-books-20210915",title:"Introducing IntechOpen Book Series - A New Publishing Format for OA Books"}]},book:{item:{type:"book",id:"8063",leadTitle:null,fullTitle:"Food Security in Africa",title:"Food Security in Africa",subtitle:null,reviewType:"peer-reviewed",abstract:"This edited volume “Food Security in Africa” is a collection of reviewed and relevant research chapters offering a comprehensive overview of recent developments in the field of food safety and availability, water issues, farming and nutrition. The book comprises single chapters authored by various researchers and edited by an expert active in the public health and food security research area. All chapters are complete in itself but united under a common research study topic. This publication aims at providing a thorough overview of the latest research efforts by international authors on Africa’s food security challenges, quality of water, small-scale farming as well as economic and social challenges that this continent is facing. Hopefully, this volume will open new possible research paths for further novel developments.",isbn:"978-1-78985-734-4",printIsbn:"978-1-78985-733-7",pdfIsbn:"978-1-83962-166-6",doi:"10.5772/intechopen.77894",price:119,priceEur:129,priceUsd:155,slug:"food-security-in-africa",numberOfPages:132,isOpenForSubmission:!1,isInWos:1,isInBkci:!1,hash:"8cbf3d662b104d19db2efc9d59249efc",bookSignature:"Barakat Mahmoud",publishedDate:"January 20th 2021",coverURL:"https://cdn.intechopen.com/books/images_new/8063.jpg",numberOfDownloads:5911,numberOfWosCitations:9,numberOfCrossrefCitations:12,numberOfCrossrefCitationsByBook:0,numberOfDimensionsCitations:19,numberOfDimensionsCitationsByBook:0,hasAltmetrics:0,numberOfTotalCitations:40,isAvailableForWebshopOrdering:!0,dateEndFirstStepPublish:"February 14th 2019",dateEndSecondStepPublish:"April 12th 2019",dateEndThirdStepPublish:"June 11th 2019",dateEndFourthStepPublish:"August 30th 2019",dateEndFifthStepPublish:"October 29th 2019",currentStepOfPublishingProcess:5,indexedIn:"1,2,3,4,5,6",editedByType:"Edited by",kuFlag:!1,featuredMarkup:null,editors:[{id:"92016",title:"Dr.",name:"Barakat",middleName:null,surname:"Mahmoud",slug:"barakat-mahmoud",fullName:"Barakat Mahmoud",profilePictureURL:"https://mts.intechopen.com/storage/users/92016/images/system/92016.jpg",biography:"Dr. Mahmoud is an international food safety expert with 30 years of experience in food safety. He was an Associate Professor in the Department of Food Science, Nutrition and Health Promotion at MSU. Prior to that, he held a postdoctoral position (in Food Safety) at Purdue University. He earned his Ph.D. in Marine Biosciences (Food Safety) from Hokkaido University (Japan). Prior to that he was a Visiting Scientist at the University of Lisbon (Portugal). He held a researcher position at the NRC (Egypt) between 1994 and 2000. He received his BSc/MSc degrees in Agricultural Sciences from Cairo University. The main goal of Dr. Mahmoud’s work is to protect public health, reduce the prevalence of foodborne illness and promote the introduction of safer food using novel technologies. Dr. Mahmoud received more than $1M from the USDA, States and Food Industry to support his research (between 2008 and 2016). Dr. Mahmoud published about 100 publications (in international journals and/or conferences), two book chapters and edited a book entitled \\'Salmonella-A Dangerous Foodborne Pathogen” (His publications have been cited more than 1200 times). He served as an editor-in-chief, editor/editorial board member for several international journals including Food Microbiology, Journal of Food Protection, Foodborne Pathogens & Disease, and African Journal of Food Science. Dr. Mahmoud has provided technical assistance in food safety in several developing countries in Africa, Asia, Central America, Middle East, and the Caribbean including Egypt, Malawi, Mozambique, Lebanon, Guatemala, the Dominican Republic, etc.",institutionString:"United States Department of Agriculture",position:null,outsideEditionCount:0,totalCites:0,totalAuthoredChapters:"0",totalChapterViews:"0",totalEditedBooks:"2",institution:null}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,coeditorOne:null,coeditorTwo:null,coeditorThree:null,coeditorFour:null,coeditorFive:null,topics:[{id:"327",title:"Food Safety",slug:"food-safety"}],chapters:[{id:"71718",title:"Understanding Africa’s Food Security Challenges",doi:"10.5772/intechopen.91773",slug:"understanding-africa-s-food-security-challenges",totalDownloads:1516,totalCrossrefCites:6,totalDimensionsCites:10,hasAltmetrics:0,abstract:"Africa, sub-Saharan Africa (SSA) in particular, has for more than 10 years recorded a steady economic growth since the advent of the new millennium. Yet, despite this stellar economic growth, it faces challenges such as rapid population growth, persistent economic inequality, climate change threats, droughts, youth unemployment, undernourishment, and food insecurity. Understanding the state of food security in Africa, and addressing the above-mentioned challenges, should be the highest priority for Africa’s Political Leadership. Not doing so will forever make Africa fail to achieve a sustainable economic development and create an inclusive shared-prosperity for its people. The African Union (AU), as well as respective national governments and regional organizations, and the international community at large, have in recent decades launched a multitude of policy initiatives aimed at addressing and tackling Africa’s food insecurity and nutrition challenges. Despite those efforts and commitments by the disparate stakeholders, much remains to be done. This chapter presents Africa’s food security and nutrition challenges, and sheds light on the climate change threats and potential consequences of the rapid population growth on Africa’s food security. The chapter concludes with policy recommendations and proposals and makes points about Africa’s bright prospects if food security were to be achieved.",signatures:"Mahamat Kabirou Dodo",downloadPdfUrl:"/chapter/pdf-download/71718",previewPdfUrl:"/chapter/pdf-preview/71718",authors:[{id:"301440",title:"Ph.D.",name:"Mahamat Kabirou",surname:"Dodo",slug:"mahamat-kabirou-dodo",fullName:"Mahamat Kabirou Dodo"}],corrections:null},{id:"67794",title:"Regime Switch and Effect on Per Capita Food Security Issues in South Africa",doi:"10.5772/intechopen.86931",slug:"regime-switch-and-effect-on-per-capita-food-security-issues-in-south-africa",totalDownloads:626,totalCrossrefCites:0,totalDimensionsCites:0,hasAltmetrics:1,abstract:"This paper examines whether the food security situation in South Africa is sensitive to the past and present governance systems. The study was aimed at reviewing the performance of key indicators: per capita land utilization, price index and consumption of a major staple food commodity (maize) in the pre- and post-apartheid periods. It also aimed at validating the application of population growth and food advocacy theories on South African food security. Time series analysis involving variables such as per capital land cultivation, consumption/tons and price/tons of maize within the period of 1970 to 2010 was conducted. Threshold autoregressive model (TAR) approach was used to capture per capita food security status of South Africans and to monitor trends under apartheid and post-apartheid eras. We found that there is a declining trend in per capita land cultivation and mixed results of per capita consumption of maize. The study revealed that population growth in South Africa has not been harnessed and there is possibility of worsening food security in the country. The long-run effect between the variables was established. The study recommends per capita targeting policy strategies for the improvement of staple food production and dietary balancing to ensure sustainable food security.",signatures:"Sunday Yiseyon Hosu and Lubabalo Qamata",downloadPdfUrl:"/chapter/pdf-download/67794",previewPdfUrl:"/chapter/pdf-preview/67794",authors:[{id:"298121",title:"Dr.",name:"Yiseyon",surname:"Hosu",slug:"yiseyon-hosu",fullName:"Yiseyon Hosu"},{id:"298126",title:"Mr.",name:"Lubabalo",surname:"Qamata",slug:"lubabalo-qamata",fullName:"Lubabalo Qamata"}],corrections:null},{id:"67624",title:"Unlocking Water Issues Towards Food Security in Africa",doi:"10.5772/intechopen.86788",slug:"unlocking-water-issues-towards-food-security-in-africa",totalDownloads:874,totalCrossrefCites:2,totalDimensionsCites:2,hasAltmetrics:1,abstract:"Water plays an important role in food security and provides the basis for healthy ecosystems and human well-being. The relationship between water and food production is key to creating resilient and sustainable food systems. This chapter will discuss the effects of water quality and scarcity with respect to food security in Africa. The effects of water availability and its usage in the African landscape and how this has impacted food security will be highlighted. Lastly, issues concerning water pollution and food safety will be tackled to identify knowledge gaps that impede food security in Africa in its efforts toward attaining the United Nations Sustainable Development Goals.",signatures:"Nokuthula Vilakazi, Kumbukani Nyirenda and Emmanuel Vellemu",downloadPdfUrl:"/chapter/pdf-download/67624",previewPdfUrl:"/chapter/pdf-preview/67624",authors:[{id:"298005",title:"Dr.",name:"Nokuthula",surname:"Vilakazi",slug:"nokuthula-vilakazi",fullName:"Nokuthula Vilakazi"},{id:"298042",title:"Dr.",name:"Kumbukani",surname:"Nyirenda",slug:"kumbukani-nyirenda",fullName:"Kumbukani Nyirenda"},{id:"298043",title:"Dr.",name:"Emmanuel",surname:"Vellemu",slug:"emmanuel-vellemu",fullName:"Emmanuel Vellemu"}],corrections:null},{id:"71000",title:"Food Safety and Food Security in the Informal Sector",doi:"10.5772/intechopen.91012",slug:"food-safety-and-food-security-in-the-informal-sector",totalDownloads:549,totalCrossrefCites:0,totalDimensionsCites:0,hasAltmetrics:0,abstract:"Food markets in the informal sector play a vital role in the livelihoods of the poor in developing countries. However, the conditions under which the informal sector operates raise concerns relating to the safety and quality of food sold. This chapter makes reference to case studies conducted by the International Livestock Research Institute’s Safe Food Fair Food project to facilitate the implementation of food safety in Sub-Saharan Africa. The purpose is to illustrate the relevance and applicability of a food chain approach in the implementation of food safety in the informal sector. Results show that some milk gets contaminated during milking, processing, cooling and handling. Such practices as adulteration and pooling were also reported to contaminate milk. The chapter concludes that it is the responsibility of all stakeholders to ensure the safety of milk along the chain.",signatures:"Bukelwa Grwambi",downloadPdfUrl:"/chapter/pdf-download/71000",previewPdfUrl:"/chapter/pdf-preview/71000",authors:[{id:"295945",title:"Ms.",name:"Bukelwa",surname:"Grwambi",slug:"bukelwa-grwambi",fullName:"Bukelwa Grwambi"}],corrections:null},{id:"72086",title:"The Role of Small-Scale Farmers in Ensuring Food Security in Africa",doi:"10.5772/intechopen.91694",slug:"the-role-of-small-scale-farmers-in-ensuring-food-security-in-africa",totalDownloads:1147,totalCrossrefCites:3,totalDimensionsCites:4,hasAltmetrics:0,abstract:"This chapter focuses on the role of African small-scale farmers in ensuring food security going into the future and the support they will need to face the projected climatic conditions. Severe climatic conditions contribute to the uncertainty in water availability for future agricultural production. Adapting to climate change and ensuring food security requires dynamic interventions that will lead to the transformation of current farming and food production patterns as well as food distribution. Nearly 95% of Africa’s agriculture is rainfed; therefore, developing and promoting rain-fed small-scale agricultural activities is a cost-effective approach for transforming rural areas in Africa and ensuring food security at local and regional levels. This is crucial in reducing vulnerability to climate change and for building sustainable livelihoods.",signatures:"Samkelisiwe Nosipho Hlophe-Ginindza and N.S. Mpandeli",downloadPdfUrl:"/chapter/pdf-download/72086",previewPdfUrl:"/chapter/pdf-preview/72086",authors:[{id:"301810",title:"Dr.",name:"Samkelisiwe Nosipho",surname:"Hlophe-Ginindza",slug:"samkelisiwe-nosipho-hlophe-ginindza",fullName:"Samkelisiwe Nosipho Hlophe-Ginindza"},{id:"308051",title:"Prof.",name:"N.S.",surname:"Mpandeli",slug:"n.s.-mpandeli",fullName:"N.S. Mpandeli"}],corrections:null},{id:"74369",title:"Food and Nutrition Security in East Africa (Rwanda, Burundi and South Sudan): Status, Challenges and Prospects",doi:"10.5772/intechopen.95037",slug:"food-and-nutrition-security-in-east-africa-rwanda-burundi-and-south-sudan-status-challenges-and-pros",totalDownloads:452,totalCrossrefCites:0,totalDimensionsCites:0,hasAltmetrics:0,abstract:"Despite receiving international technical assistance over many years, achieving food and nutrition security has remained elusive for many developing countries. Low technological capability, inefficient production systems, increasing populations and lately climate variability, affect food production, leading to stagnation or modest gains in food and nutrition security in many nations. For many African countries, food and nutrition security continues to improve, despite the slow pace. In the East African Community, political stability, ambitious economic planning, the quest for higher agricultural productivity, improving educational achievement, sanitation and health, are contributing to improving food and nutrition security. To support the process, Rwanda, established Vision 2020, while Burundi and South Sudan have yet to develop plans for a coherent development blueprint. The blue prints of the Member States bore Vision 2050 for the East African Community and Vision 2063 for the African Union. This chapter examines the status of food and nutrition security in Rwanda, Burundi and South Sudan. It gives country-specific recommendations for achieving it-including investment in agriculture and agribusiness, value addition of agricultural commodities, decelerating population growth, using adaptive research to solve farmer-problems, strengthening farmer-organizations and integrating variables that influence food and nutrition security achievement.",signatures:"Michael N.I. Lokuruka",downloadPdfUrl:"/chapter/pdf-download/74369",previewPdfUrl:"/chapter/pdf-preview/74369",authors:[{id:"306514",title:"Associate Prof.",name:"Michael N.I.",surname:"Lokuruka",slug:"michael-n.i.-lokuruka",fullName:"Michael N.I. Lokuruka"}],corrections:null},{id:"74388",title:"Food and Nutrition Security in East Africa (Kenya, Uganda and Tanzania): Status, Challenges and Prospects",doi:"10.5772/intechopen.95036",slug:"food-and-nutrition-security-in-east-africa-kenya-uganda-and-tanzania-status-challenges-and-prospects",totalDownloads:750,totalCrossrefCites:1,totalDimensionsCites:3,hasAltmetrics:0,abstract:"Achieving food and nutrition security remains a tall order for developing countries. The FAO, IFPRI, WFP, UNICEF and other international bodies continue to provide active support in order to achieve global food and nutrition security. However, low technological capability, inefficient production, insignificant economic growth, increasing populations and lately climate variability, affect food production, leading to either stagnation or modest gains in food and nutrition security in different regions of the World. For African countries, food and nutrition security continues to improve, albeit at a slow pace, although the recent breakout of COVID-19 is bound to lead to a decline in food production, in the short and mid-term. In the East African Community, political stability, ambitious economic planning, the quest for higher agricultural productivity, improving educational achievement, improving sanitation and health, are contributing to the improving food and nutrition security. To hasten the process, Kenya, Uganda and Tanzania embraced Vision 2030, Vision 2040 and Vision 2025, respectively. These grand, socio-economic plans bore Vision 2050 in the East African Community and Vision 2063 for the African Union. This chapter examines food and nutrition security in Kenya, Uganda and Tanzania, and provides country-specific recommendations for achieving it. These include investing in agriculture, decelerating population growth, using adaptive research to solve farmer-problems, strengthening farmer-organizations and the formation of cooperatives.",signatures:"Michael N.I. Lokuruka",downloadPdfUrl:"/chapter/pdf-download/74388",previewPdfUrl:"/chapter/pdf-preview/74388",authors:[{id:"306514",title:"Associate Prof.",name:"Michael N.I.",surname:"Lokuruka",slug:"michael-n.i.-lokuruka",fullName:"Michael N.I. Lokuruka"}],corrections:null}],productType:{id:"1",title:"Edited Volume",chapterContentType:"chapter",authoredCaption:"Edited by"},subseries:null,tags:null},relatedBooks:[{type:"book",id:"799",title:"Salmonella",subtitle:"A Dangerous Foodborne Pathogen",isOpenForSubmission:!1,hash:"ba452d8a24ef16b1267d2854b28f6e6a",slug:"salmonella-a-dangerous-foodborne-pathogen",bookSignature:"Barakat S. M. 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2017",dateReviewed:"May 4th 2018",datePrePublished:null,datePublished:"January 30th 2019",book:{id:"6671",title:"Paint and Coatings Industry",subtitle:null,fullTitle:"Paint and Coatings Industry",slug:"paint-and-coatings-industry",publishedDate:"January 30th 2019",bookSignature:"Faris Yilmaz",coverURL:"https://cdn.intechopen.com/books/images_new/6671.jpg",licenceType:"CC BY 3.0",editedByType:"Edited by",editors:[{id:"36900",title:"Dr.",name:"Faris",middleName:"Sad",surname:"Yılmaz",slug:"faris-yilmaz",fullName:"Faris Yılmaz"}],productType:{id:"1",title:"Edited Volume",chapterContentType:"chapter",authoredCaption:"Edited by"}},authors:[{id:"186371",title:"Associate Prof.",name:"Imed",middleName:null,surname:"Gargouri",fullName:"Imed Gargouri",slug:"imed-gargouri",email:"imed.gargouri@fmsf.rnu.tn",position:null,institution:{name:"University of 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research that has attracted the attention of many scientists over the past few decades. Municipal solid waste, which has traditionally been studied the most, is a stream that, by weight, is at relatively low percentages compared to the total amounts of solid wastes generated worldwide. Emphasis is lately given on special solid waste streams that may, or may not, have hazardous properties. The aspect of sustainability has been particularly introduced into solid waste management during the past two decades. For example, “sustainable landfilling” was a term introduced during the nineties to suggest the use of stabilization techniques before landfilling to reduce landfill emissions.
\r\n\r\n\tThis book, therefore, intends to provide the reader with a comprehensive overview of the current trends of research that aims to focus on this diverse nature of sustainable solid waste management: from valorization techniques to legal, economic, and behavioral aspects and from LCA to quality assessment methods.
",isbn:"978-1-80356-327-5",printIsbn:"978-1-80356-326-8",pdfIsbn:"978-1-80356-328-2",doi:null,price:0,priceEur:0,priceUsd:0,slug:null,numberOfPages:0,isOpenForSubmission:!1,isSalesforceBook:!1,hash:"e3e2cbc06fea6858df1f375d49431b66",bookSignature:"Prof. Suhaiza Zailani",publishedDate:null,coverURL:"https://cdn.intechopen.com/books/images_new/11530.jpg",keywords:"Solid Waste Management, Sustainable, Supply Chain, UN Sustainable Development Goals, Venture, Solid Waste Management, Circular Economy, Sustainable Solid Waste, Behavioral Insights, Behavioral Aspects, Solid Waste Management, Business Sustainability",numberOfDownloads:null,numberOfWosCitations:0,numberOfCrossrefCitations:null,numberOfDimensionsCitations:null,numberOfTotalCitations:null,isAvailableForWebshopOrdering:!0,dateEndFirstStepPublish:"February 16th 2022",dateEndSecondStepPublish:"March 16th 2022",dateEndThirdStepPublish:"May 15th 2022",dateEndFourthStepPublish:"August 3rd 2022",dateEndFifthStepPublish:"October 2nd 2022",remainingDaysToSecondStep:"2 months",secondStepPassed:!0,currentStepOfPublishingProcess:4,editedByType:null,kuFlag:!1,biosketch:"Dr. Zailani was awarded the UM Distinguished Researcher in 2016, Emerald Literati Network Awards for Excellence in 2016, and last year, and was listed as one of the recipients for the Top 80 UM Researchers amongst the World’s Top 2% of scientists by Sandford University.",coeditorOneBiosketch:null,coeditorTwoBiosketch:null,coeditorThreeBiosketch:null,coeditorFourBiosketch:null,coeditorFiveBiosketch:null,editors:[{id:"172845",title:"Prof.",name:"Suhaiza",middleName:null,surname:"Zailani",slug:"suhaiza-zailani",fullName:"Suhaiza Zailani",profilePictureURL:"https://mts.intechopen.com/storage/users/172845/images/system/172845.png",biography:"Suhaiza Zailani is a Professor of Supply Chain with the Faculty of Business and Accountancy, University Malaya. 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From chapter submission and review to approval and revision, copyediting and design, until final publication, I work closely with authors and editors to ensure a simple and easy publishing process. I maintain constant and effective communication with authors, editors and reviewers, which allows for a level of personal support that enables contributors to fully commit and concentrate on the chapters they are writing, editing, or reviewing. I assist authors in the preparation of their full chapter submissions and track important deadlines and ensure they are met. I help to coordinate internal processes such as linguistic review, and monitor the technical aspects of the process. As an ASM I am also involved in the acquisition of editors. 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Hypersensitivity of visceral mechanoreceptors could result from excessive production of modulatory neurotransmitters. In addition to direct stimulation of stretch-activated channels on primary afferent neurons located in dorsal root ganglia (DRG), chemicals produced by different target cells (such as smooth muscle cells and interstitial cells of Cajal) in response to stretch or inflammation play an important role in the neuromodulation of nociception. The incidence of persistent, episodic, or chronic visceral pain is more prevalent in females, which also suggests hormonal regulation. Despite extensive research of the properties of pelvic and splanchnic afferent nerves, little is known about the mechanisms underlying normal and pathological signal transduction pathways underlying many functional diseases. Despite considerable efforts by the scientific community and the pharmaceutical industry to develop novel pharmacological treatments aimed at chronic visceral pain, the traditional approaches to identifying and evaluating novel drugs for this target have largely failed to translate into effective IBS treatments [1]. A better understanding of these processes has direct implications for the development of more effective therapies. During the last decade, we identified that DRG neurons can be affected by ATP, NO, estrogen, and other mediators producing neuronal hyper- or desensitization that may unravel the enigma of the development of chronic pelvic pain associated with IBS. Moreover, our recent data that estrogen can gate primary afferent response to modulate nociception support the idea about involvement of peripheral central system in etiology of a wide range of the functional and inflammatory gastrointestinal diseases [2].
\nPelvic nerve afferent fibers innervating the visceral organs of the lower colon have been well characterized (reviewed in Ref. [3]). In general, during colonic distension, a large number of pelvic EPANs show static levels of discharge. Stretches that lead to the opening of stretch-activated (SA) channels on the plasma membrane lead to the selective or nonselective opening of different cation and anion channels in nodose ganglia and DRG neurons. Thus, depending on the cell type and channel type, EPANs activation may result in hyperpolarization, depolarization, or primarily Ca2+ influx. The function of SA channels in the plasma membrane differs between various cell types. Influx of Ca2+ may repolarize the plasma membrane via activation of Kca channels and inactivating of voltage-gated calcium channels (VGCCs), and thus influence adaptation rates of sensory neurons during ongoing stimulations.
\nThe cell bodies of primary visceral spinal afferent neurons are located in the lumbosacral (L1-S1) DRGs that transmit information about chemical or mechanical stimulation from the periphery to the spinal cord. Nociceptors are small- to medium-size DRG neurons whose peripheral processes detect potentially damaging physical and chemical stimuli. Until recently, ATP release from nonneuronal cells was thought to be exclusively as result of injury. It is now clear that certain integral membrane proteins contain an ATP-binding cassette so this neurotransmitter can act as signaling molecule modulating sensory afferent nerve terminals. Six P2X receptors are expressed in DRGs. Significantly, the P2X3 receptor is found exclusively in a subset of small diameter capsaicin sensitive peripheral sensory neurons (presumably nociceptors) [4]. Today, multiple lines of evidence suggest that ATP signaling via P2X receptors contribute to different pain phenotypes, therefore P2X antagonists may be useful analgesics. The availability of P2X receptor-specific antagonists also holds the promise of revealing the cellular and molecular neurobiology underlying pain states underlying functional diseases [5]. With sufficiently high levels of ATP, P2X and SA channels (which has a greater permeability for Ca2+ than Na+) would depolarize nerve terminals directly producing action potentials and leading to sensation of pain. On the other hand, the response of EPANs may be tonically inhibited by NO produced by peripheral nerve terminals. The peripheral sensitization of nerve fibers is transient depending on the duration of stimuli and the presence of visceral (colonic) inflammation.
\nChanges in pain perception and variations of symptoms throughout the menstrual cycle, as well as sexual intercourse triggering symptoms in a significant portion of females diagnosed with irritable bowel syndrome (IBS), painful bladder syndrome (PBS), chronic pelvic pain (CPP), and other function syndromes, points to a connection with sex steroids. Several lines of evidence indicate that 17β-estradiol (E2) directly influence the functions of primary afferent neurons. Both subtypes of estrogen receptors (ERα and ERβ) are present in DRG neurons including the small-diameter putative nociceptors [4]. In vitro, ATP-sensitive DRG neurons respond to E2 [6, 7], which correlated well with the idea that visceral afferents are E2 sensitive: (i) visceral pain is affected by hormonal level in cycling females; (ii) there are sex differences in the prevalence of functional disorders involving the viscera; and (iii) putative visceral afferents fit into the population of DRG neurons that are sensitive to E2 [7]. These data suggest that in addition to the CNS actions, E2 can act in the periphery to modulate nociception [6, 7]. E2 modulates cellular activity by altering ion channel opening, G-protein signaling, and activation of trophic factor-like signal transduction pathways. These effects have been ascribed to membrane-associated receptors [8]. The results from our laboratory and others indicate that E2 acts in DRG neurons to modulate L-type VGCC and through group II metabotropic glutamate receptors [6].
\nE2 has a significant role in modulating visceral sensitivity, indicating that E2 alterations in sensory processing may underlie sex-based differences in functional pain symptoms [9]. Indeed in most clinical studies, women report more severe pain levels, more frequent pain, and longer duration of pain than men [10]. Little is known about E2-mediated mechanisms in peripheral nervous system, but the fact that DRG neurons express ERs and respond to E2 treatment suggest that they are a potential target for mediating nociception. E2 modulation of nociceptive response depends on the type of pain, its durations, and the involvement of other nociceptive-mediated mechanisms.
\nE2 (both short-term and long-term exposure) significantly decreased the nociceptive signaling in viscerally labeled DRG neurons [6, 7]. Thus, in addition to central regulation, estrogen may affect nociception associated with IBS peripherally.
\nMost of the current literature pertains to specific functional syndromes defined by medical subspecialties. These include: IBS (gastroenterology); CPP (gynecology); PBS (urology); fibromyalgia (rheumatology); and others. Many reports described the substantial overlaps between two or more of these syndromes [11, 12]. Moreover, clinical presentations of functional syndromes lack a specific pathology in the affected organ but may respond to a viscero-visceral cross-sensitization in which increased nociceptive input from an inflamed organ (i.e., uterus) sensitizes neurons that receive convergent input from an unaffected organ (i.e., colon or bladder). The site of visceral cross-sensitivity is unknown.
\nRecent studies from our laboratory demonstrated that hormonal modulation of visceral inputs of primary afferent nociceptors located in the dorsal root ganglia (DRG) is responsible for changes observed in the perception of pain during the etiology of functional pain syndromes [2]. Individuals suffering from CPP frequently have pain emanating from several visceral organs. Viscero-somatic and viscero-visceral hyperalgesia and allodynia lead to the perception of pain spreading from an initial site to adjacent areas. Patients with CPP may at first have only one source of pain in the pelvis, but numerous mechanisms involving the central and peripheral nervous systems may result in the development of painful sensations in adjacent organs, such as IBS being associated with lower colonic pain.
\nSimilar to other chronic diseases, a multicomponent conceptual model of IBS, which involves physiologic, cognitive and behavior factors will be necessary for developing new therapies. The different systems such as neuroendocrine regulation, pain modulation, and autonomic response will affect ascending aminergic systems (\nFigure 1\n).
\nDifferent regulatory mechanisms involved in irritable bowel syndrome.
From a public health perspective, a substantial impact on our knowledge of nociceptive diseases such IBS will help achieve a deeper understanding of data presented in clinical aspects of these symptoms. Only a thorough understanding of the mechanism implicated in these phenomena can truly contribute to the designing of new and more efficient therapies.
\nEnzyme linked immunosorbent assay (ELISA) has existed for 50 years and ELISAs with different technical solutions are still being developed, which improves and expands the range of application.
The test was first described by Engvall and Perlmann in 1971 [1, 2, 3] and was based on the work of Avrameas, who used enzyme linked antibodies in histochemistry [4, 5]. The method was quickly developed for sero-diagnosis of trichinosis [6] and antibodies to
Since the discovery there have been numerous applications of ELISA, used to detect both antigens and antibodies. Besides the detection of protein antigens ELISAs that permit the determination of antibodies to native and denatured DNA [8, 9], polysaccharide antigens [10, 11, 12] and phospholipids [13] have been optimized. In fact, sometimes the name ELISA is applied to tests in which there are no antibodies, but instead specific protein–protein interactions are used. From the perspective of optimization, validation and standardization such tests can be treated in the same way. Regarding protein antigens the sensitivity of ELISA is usually in the pg/ml range [14].
When developing a diagnostic test, precise and optimal performance conditions must be found for all the steps within the test protocol. This ensures that the entire procedure is optimal. Before routine usage in diagnostics, for example, the newly developed, or a newly modified procedure must be proven to be accurate, precise and reproducible. Also, in order to measure the values obtained with the test, it is necessary to standardize the test. Therefore, optimization, validation and standardization (OVS) of ELISA are extremely important and necessary, especially if it is to be used in clinical or veterinary medicine. This chapter will present the procedures by which ELISA is characterized in an understandable and precise way.
Reviewing the literature, we noticed that the described boundaries between optimization, standardization and validation are not clear enough. The reason for this is that in certain situations performing a single ELISA can lead to a completion of both validation and optimization characteristics, which is completely valid. Before going into more details and in order to avoid confusion it is suitable to clearly define these three terms.
According to Merriam-Webster dictionary,
ELISA most often serves to measure the presence or quantity of antibodies or antigens, or biomolecules in general which can be recognized by antibodies. In biological matrices (such as serum, plasma, blood, urine and saliva) ELISA is an important diagnostic tool used to detect various antigens and antibodies. Indirect or direct ELISAs are used in medical product development, particularly for testing vaccines and new drugs. ELISA with specific antibodies can be designed to measure impurities within the medical products resulting from the production process. Antibody assays against these impurities should also be developed and validated for testing the levels of the impurities, which should be kept at a minimum in order to avoid adverse immune responses. For immunogenic substances with expected low concentrations, such as cytokines, hormones, toxins etc., sandwich ELISA is used.
Irrespective of the ELISA design (indirect, direct or sandwich), OVS principles are the same. Of paramount importance for any bioanalytical method is that it is well characterized, fully validated and documented to a satisfactory standard in order to yield reliable results.
The first step in ELISA development is optimisation, which is followed by standardization and finaly validation.
Optimization of an ELISA is essential to its success. Since ELISA is a multistep procedure, each component can be individually tested prior to the start of an experiment.
ELISA procedure consists of antigen or antibody coating, saturation, analyte application, detection with appropriate antibodies, primary or secondary and signal detection. Between each step the plate is washed. A variety of samples can be tested with ELISA, and the choice of assay conditions will depend upon the complexity of the sample and the expected amount of analyte present. Optimization is the establishment of ideal concentrations of each assay reagent and ideal conditions for each step and that must be done empirically. The cornersotne of any ELISA is the selection of the protocol type: direct, indirect or sandwich; which is dependent on the type of sample, avaliable reagents and the concentration of the analyte, keeping in mind that the procedure should be as straight forward as possible.
Numerous factors should be tested, such as the concentration of antigen, or antibody used for coating, temperature, the duration of individual steps the type of coating buffer, such as phosphate-buffered saline (PBS) or carbonate buffer, sample preparation methods (with or without EDTA, decomplementation, serum or plasma or whole samples). Plate saturation is also a step which requires optimization such as different concentration of bovine serum albumine (BSA), nonfat-dried milk, or whole serum from different animals. Here we will discuss the most important steps of the optimization procedure.
The first step in ELISA is coating wells with antigen or capturing antibodies. Most often this consists of applying a protein solution in PBS or carbonate buffer to microttiter plate wells. The microtiter plates for coating with proteins are special plates with modified surface, i.e. highly charged polystyrene surface with high affinity to molecules with polar or hydrophilic groups. This kind of surface has a high binding capacity for proteins, including globular antibodies and ensures proper antibody orientation. On the other hand ELISA for lipid antigens is performed on a hydrophobic surface, suited for non-protein antigens, which are not soluble in PBS or carbonate buffer, but are dissolved in an apropriate alcohol. Irrespective of the type of antigen the whole surface of the well bottom must be covered. If the whole surface is not covered the absorbance read will be lower, and if excess antibody/antigen is present, layers of antibody/antigen may form and wash away in subsequent steps, which again leads to lower signal. Figure 1 shows the dependance of absorbance on the ammount of antibody/antigen used for coating. For the optimized protocol it is important to select that antigen/antibody concentraion that gives the highest absorbance, marked with a red circle in Figure 1, which ensures that the complete well surface availiable for binding is covered in a monolayer. This principle should be followed regardless of the type of antigen/antibody or the ELISA type. For axample, in sandwith ELISA the wells are covered with capture antibodies, either whole IgG or Fab fragments and in direct and indirect ELISA with the antigens.
Dependance of absorbance on the ammount of antibody/antigen used for well coating in ELISA.
The process of coating an ELISA plate with antigen relies on the binding activity of the solid phase of the well, which immobilizes biomolecules on the well surface. Step after that must be blocking. During blocking free binding sites at the bottom of the wells become saturated with a blocking buffer in order to prevent the possibility of nonspecific binding and the residual binding capacity of the wells, thus greatly improving the signal-to-noise ratio and specificity. Without appropriate blocking the detection antibody could bind nonspecifically alongside the antigen, resulting in high background signal and low sensitivity.
There is a variety of blocking buffers, to choose from, not one of which is ideal for every situation. Although these buffers are called blocking buffers they usually contain a blocking component such as BSA, nonfat-dried milk, casein or whole serum. Every blocking buffer represents a compromise between reducing the background and maintaining specificity. Whole sera and serum protein albumin can cause non-specific ELISA signals in certain circumstances [15].
Even different BSA preparations show variations in the blocking activity of non-specific binding in ELISA. To prevent false positive results from cross reactive antibodies or non-specific binding of ELISA reagents to BSA, alternative blocking agents can be used and even no protein can be included in the blocking buffer [1]. These different blocking agents, (as well as their different concentration, incubation time, etc) should be tested in parallel, to discover the best way of saturation for each individual ELISA system.
It is almost always necessary to dilute samples for ELISA test, so the choice of the diluent is important. Generally, standard diluent should be as similar as possible to the matrix of the sample. For example, PBS with BSA is a good serum replacement in ELISA and is most often used for biologycal samples. The next important diluent component is non-ionic detergent (Tween 20, Triton X-100, CHAPS) that, in low concentrations, prevents non-specific (hydrophobic) protein–protein interactions. The specific binding is usually more resistant to the detergent. Detergents in one step do not provide a permanent barrier to biomolecule non-specific attachment in the following steps because it washes away with water or aqueous buffer, so in certain situations, detergents should be present in all the diluents/buffers.
It may be necessary to choose a different diluent than PBS/Tween/BSA, if the analyte is not serum. In that case, it is necessary to check the standard curve and linearity of dilution for the experimantal sample. The reason for this is the influence of the components of a standard diluent or matix on antigen/antibody interactions. In such cases spike-and-recovery or linearity-of-dilution experiments should be performed.
The goal in assay development is to achieve high signal-to-noise ratio while maintaining optimal responses. The sample matrix may contain interfeering components that affect assay response to the analyte by introducing a difference in comparison to the standard diluent. In order to asses this phenomenon, spike-and-recovery experiment is designed.
The idea of spike-and-recovery is that you add (spike) a certain amount of standard into the sample buffer or the samples, and measure them in parallel with samples with no standard added. Sometimes one can compare the same amount of analyte added into the natural test sample matrix and identical spike added to the standard diluent. So it can be seen whether you can measure (recover) the exact amount again, and how much you can recover from it in percentages. If, for any reason, you can not recover the same amount in comparison to a control, this means that something in the test solution is not in favor of the assay, so one should proceed with finding the right standard diluent.
Linearity-of-dilution experiments provide information about the precision of the assay results for different diluted samples in the chosen sample diluent. These experiments are performed to demonstrate that highly concentrated samples can be accurately measured by diluting into the assay’s quantitative range and the concentration can be calculated by multiplying the measured concentration by the dilution factor. Linearity-of-dilution experiment in practise means the measurement of at least three dilutions in the appropriate range in the selected diluent. There are two different ways to perform a linearity-of-dilution experiment, both with the same outcome. The usual method implies using a highly concentrated sample and then testing several different dilutions of that sample in the chosen sample diluent. Alternatively one can first prepare several different dilutions of a low concentration sample and then spike it with the same amount of the analyte before testing. If a sample does not exhibit linear dilution (i.e. linear dependence of absorbance on dilution), the situation can be that one has missed the range of linearity, as generally speaking linearity rarely or never exists over the entire range of concentrations; or that the matrix component is interfering with the measurement at the given dilution. Sometimes, matrix interference occurs if an interfering factor is present at concentrations above a certain threshold, and when the sample is diluted, interference is no longer observed. This kind of testing of a novel bioanalytical method is required by the EMA [16, 17].
When testing an experimental sample it is important to test several dilutions, all in duplicate or triplicate in conjunction with a known standard to ensure that the final results fall within the linear portion of the standard curve. This ensures the accuracy of the result. In highly concentrated samples underestimation of the concentration can occur, while in highly diluted samples overestimation can occur. Prepare different concentrations of the sample, keeping in mind the detection limit of the substrate. At this point, it is very suitable to detect maximal quantity of sample that can be detected, that is the last concentration after which there is no further absorbance increases (the same principle as for antigen coating optimization), Figure 1. This way the upper limit of the method is determined which enables the optimization of the next step.
At this point of optimisation, if sample is sera, high unspecific absorbance can occur, which is not related to the concentration of the sample/analyte. This can occur if the sera is not decomplemented, because active complement binds to antibody Fc. Heat-inactivation of serum for 30 minutes at 56°C eliminates complement activity, but one must keep in mind that different immunoglobulin isotypes and immunologbulins from different species show different sensitivity to heat treatment [18]. So, it is important to carefully consider or test the inactivation step.
ELISA is largely dependent on the choice of antibodies used, so antibodies should be carefully chosen. Based on the type of sample and the expected analyte concentration, the choice of monoclonal or polyclonal antibodies, or even the combination of both, should provide optimal signal-to-noise ratio [19]. Each antibody type offers distinct advantages.
The interaction between antibodies and their antigens is described by specificity, affinity, and avidity.
Specificity is an indication of whether an antibody binds solely to a unique epitope from a single antigen in a single species, or whether it binds to similar epitopes present on several molecules from the same or a few different species, i.e. whether it is cross-reactive. Specificity is the most important quality of an antibody, and this is the principle that ELISA is based upon, so a carefull selection should be made.
Affinity describes the strength of binding of an antibody with an antigen. This binding is a reversible interaction and affinity determines how much antigen is bound by an antibody at any particular moment, which is dependent upon how quickly this binding occurs, and for how long the interaction lasts. High affinity antibodies should be used in all types of immunoassay because they rapidly produce a large number of stable interactions and provide the most sensitive detection.
Avidity is a less intuitive term than affinity as it is based on affinity, but is highly influenced by the the total number of antigen binding sites or valency, which determines the overall stability of the antibody–antigen interaction. Therefore, avidity varies with antibody isotype and whether it is intact or fragmented. Additional factors which determines avidity are the structure of the antibody, the length and motility in the hinge region and the space between the Fab fragments.
When available, one should always choose monoclonal antibodies over polyclonal antibodies, in fact, commercial ELISA kits almost always utilize monoclonal antibodies. Monoclonal antibodies have specificity for a single epitope, usually a small part of the antigens’ surface. Monoclonal antibodies are therefore less likely to interact with closely-related proteins and are not generally expected to trigger non-specific signals in an immunoassay. Polyclonal antibodies are a mixture of antibodies with increased specificity to the antigen, therefore they bind different epitopes. Commercial polyclonal antibodies are often affinity purified or cross-adsorbed, but still the posibility of crossreactivity is higher. In addition, polyclonal antibody preparations can show batch to batch variations which should not be the case with monoclonal antibodies.
The advantage of using polyclonal antibodies is that they rarely fail to bind to the antigen due to a single blocked antibody binding site, antigen configuration change, or misfolding, although the latter are more important in tests other than ELISA. When combining monoclonal antibodies as in sandwich ELISA it is important to check literature or to test experimentaly the compatibility of the antibodies in terms that they do not share an epitope or for steric hinderance. Matched pairs are the basis of many sandwich ELISAs, either in kits or for in house assay set up. Matched antibody pairs means they are capable of detecting different epitopes on the same protein antigen, so they can be used together in a sandwich ELISA.
Sometimes the ELISA sensitivity can be increased by using indirect detection with polyclonal antibodies instead of direct detection with a monoclonal antibody, due to higher levels of polyclonal antibody binding to the target antigen. For cost reduction it can also be the combination of monoclonal capture with polyclonal detection.
After careful antibody selection, serial dilutions of capture antibodies should be carefully prepared for proper titration of antibody concentration. This is performed according to the previously mentioned principle of detecting maximum ammount of the component (in this case detection antibody) after which there is no further absorbance increase, Figure 1. Again, the ideal concentration should provide the highest signal and lowest noise.
As ELISA is a method which basicaly consists of overlaying different components which specificaly interact in each step (except washing) an optimization is required which follows the principle of titration until the complete coverage of the previous layer. Often the enzyme conjugate, i.e. enzyme responsible for color development, is already chemically bound to the detecting antibody, thereby enabling its direct use as a detection antibody in immunoassays. If this is not the case then enzyme concentration should be optimized too.
In this step, the first point is choosing the apropriate enzyme conjugate, depending on the needs of the researcher. The enzymes should be stable at typical assay temperatures: 4°C, 25°C, and 37°C; have a shelf life greater than six months when stored at 4°C; be inexpensive and commercially available. The enzymes should also survive the necessary conjugation conditions and yield productive conjugation. The enzymes should have an easily measurable activity; with high substrate turnover number. Horse radish peroxidase (HRP) and calf intestine alkalne phosphatase (ALP) are two most widely used enzymes for detection in ELISA assays [20]. HRP is usually conjugated to an antibody in a 4:1 ratio. For ALP the ratio is a little more unfavorable, 2:1, but the conjugate is more stable [21]. These enzymes are typically used because they each meet most, if not all, of the criteria necessary to produce a sensitive, inexpensive, and easily performed assay.
All enzyme-linked immunoassays, imply the usage of the enzyme substrate. Colorimetric ELISAs usually require soluble colored reaction products. The decision which substrate to choose depends on the desired sensitivity, reaction time, and the detection device. For colorimetric detection the most desirable substrates quickly produce intensely colored reaction products. When the analyte amounts span a wide range of concentrations (large dynamic range), then it is more suitable to use substrates that produce color over a longer time period (15 to 30 minutes) because then, one is able to detect the wider range of analyte-dependent color intensities. For assays with a timed endpoint, the reaction is stopped with an inhibitor suitable for the specific enzyme substrate combination after a defined time period that stops further color development. This allows detection to be performed within a reasonable time; for this, a substrate that has a “slow” reaction rate (15 to 30 minutes to completion) is optimal.
Both HRP and ALP have substrates that yield soluble colored reaction products.
The most common substrates that produce soluble reaction products with HRP are: TMB (3,3′,5,5′-Tetramethylbenzidine), ABTS (2,2′-azino-di[3-ethylbenzthiazoline] sulfonate), and OPD (o-phenylenediamine). TMB is a highly sensitive substrate, safe for laboratory workers. Due to its rapid reaction rate, it is ideally suited for on-line kinetic analysis. TMB can also be used in endpoint assays by stopping the reaction with 1 M phosphoric acid. ABTS is considered an all-purpose substrate. Although it is less sensitive than either TMB or OPD, it has the widest working range of any substrate currently available for peroxidase or alkaline phosphatase. Its reaction rate is suitable for endpoint assays and is easily stopped with 1% SDS (sodium dodecyl sulfate), which does not change the color or the absorbance of the reaction product. OPD was once the most popular substrate for peroxidase. It is slightly less sensitive than TMB, but it is cancerogenic.
The most commonly used substrate that produces a soluble reaction product with ALP is p-NPP (p-nitrophenylphosphate). pNPP is a substrate with a low reaction rate, so it usually takes 30 to 60 minutes for the dye to develop optimally. This property makes it possible to increase the sensitivity by increasing the reaction time period. At the same time, this property makes the pNPP substrate unsuitable for kinetic analysis [22].
Factors that affect the measurement of enzymatic activity are temperature, buffer composition (pH, ionic strength), build-up of product inhibitors, the increase in back-reaction as the product concentration increases, stability of the enzyme and sometimes exposure to light. As most of these facors such as pH and substrate depletion, are known, commercially available reagents are optimized for composition and concentration in order to control these parameters. For novel ELISA optimization of the most concern are reaction time and temperature.
If the antigen can clearly be detected then the substrate is appropriate. If the antigen is below the threshold for detection then one should select a more sensitive substrate.
It should be noted that the detection methodologies for ELISA are few, but the most prevalent in the laboratories is colorimetric. In addition, fluorescent and luminescent are also used.
In colorimetric detection the amount of color in each well is read by a spectrophotometer and samples are compared relative to one another or with the use of a standard curve derived from known analyte concentrations.
Fluorescent substrates [23] for ALP and HRP can potentially yield a higher signal, leading to increased sensitivity and broader dynamic range. This kind of detection requires black plates, which are also availiable with various degrees of hydrophobicity and a fluorescent plate reader is required. Fluorescence yielding substrates have a shorter half-life than colorimetric substrates, so the signal is declining over time. This kind of ELISA is useful for measuring immune responses because of broader dynamic range [19].
The same detection antibodies conjugated with ALP or HRP, can also be used for chemiluminescent assays [24]. In this type of experiment, ALP, for example, will modify a substrate, forming a chemiluminiscent product which creates light emission. ALP chemiluminiscent substrates can have pg/ml sensitivity. The signal can be read in black or white opaque ELISA plates and a luminometer is required. The advantages of this detection type are typically a higher dynamic range and lower background signal. The signal is not as stable as the colorimetric or fluorescent detection and must be read within a short time of generating the signal.
The type of substrate used depends on several factors, most notably the desired assay sensitivity and signal to bakground ratio.
Many laboratories have independently developed ELISA techniques for their own purposes. For results to be valid they must be comparable with results of the same ELISA test performed in different laboratories. Consistency in the assessment of ELISA results in different areas of application (diagnostics, production control, scientific research, immunogenicity assessment etc.) requires standardized and acknowledged methodological protocols. Protocol harmonization progress with respect to the international standardization and validation of this technique has been made.
Today, leading regulatory agencies for specific guidance on immunogenicity assessment of biotherapeutic products are part of EMA and WHO, [25] and there are other agencies. The National Institute for Biological Standards and Control (NIBSC), for example, part of UK Medicines and Healthcare products Regulatory Agency (MHRA), is of great importance to the field of biological standardization. It produces over 90% of the biological international standards in use around the world. The WHOs’ Biological Reference Materials are established through a standard procedure, [26] in which representative materials are tested by participating laboratories using their own methodologies and coordinated by a responsible WHO Collaborating Center [27]. Upon establishment of the reference preparation by the Expert Committee on Biological Standardization (ECBS), the material is assigned a unitage and serves as the comparator against which results from laboratories can be standardized and compared, irrespective of the location or the methods employed. This enables the results of bioanalytical methods, including ELISA, to be comparable. Based on international standards „ working standard” (i.e. in-house or secondary standards) are evaluated and compared, and subsequently adequately used.
At first glance, it is very simple to explain the process, i.e. the term of standardization in ELISA: comparing the absorbance of a sample with the absorbance of the known concentration of the standard (in-house or commercial) and based on that, determining the unknown concentration.
If the ELISA is intended for the measurement of the final detectable dilution, as in titration experiments, and not for the measurement of biomolecule quantity a reference standard may not exist.
Then the need exists for establishing a reference standard. For any ELISA, consideration must be given to the selection of standards which represent, on average, what would be expected of an immune response of the organism in question. Immunogenicity assessment relies on the measurement of antigen induced antibodies in serum or plasma. Such antibodies are heterogeneous in terms of classes, subclasses and alotypes, concentration as well as antigenic specificity. Some will neutralize the biological activity of the antigen, others will not, despite the high affinity/avidity. Irrespective of the type of ELISA system used, endpoint titration is a function of both antibody concentration and avidity. And finally, as every sample is unique with vast individual differences among humans, for example, it is not possible to make a straightforward comparison with standard antibodies. Nevertheless, although the ideal is unreachable, if wanting to produce valid and reproducible data a reference standard must be established.
The physical quantity to be measured in ELISA is absorbance. Absorbance is influenced by test parameters and photometric instrumentation, so raw, corrected or normalized OD values [28] cannot be used for inter-laboratory standardization. This is why end-point titration or determination of highest serial dilution which demonstrates a minimum of antibody activity is often used for measuring the immune response in diagnostics and vaccinology. Under some circumstances, quantitative data are not required for diagnostic purposes and sometimes end-point titration is sufficient, with an adequate semi-quantitative standard. End-point titrations are labor-intensive, costly and impractical for most routine diagnostic purposes.
In order to overcome the relativity of the measured absorbance a notion of “percent positivity” (PP) is accepted, this way the absorbance of each sample tested is expressed as a percentage of a highly positive reference standard. Although semi-quantitative, PP is expressed on a continuous scale of 0–100 and has two major advantages, first, it requires only a single dilution and second, it does not assume parallelism or uniform background activity. Therefore, it may be used for inter-laboratory standardization.
Even with measurements with qualitative standard curve, it is not correct to determine the result from a single sample dilution measurement. This can only be acceptable if there is a parallelism in dilution curves between the sample and the standard. If more quantitative data are needed, PP values can be converted to units which are directly proportional to antibody activity.
Sometimes an elegant and appropriate way to quantify samples is competitive or inhibitory ELISA. When performing competitive ELISA, one applies the sample preincubated with the same antigen used for plate coating and measures the amount of non inhibited antibodies. There is a negative relationship between color intensity and the amount of test sample antibody inhibited by antigens. Percent inhibition (PI) of the color produced by the standard competing antibody is more widely used. The development of consistent standard curves for this kind of assay is extremely difficult, but still possible.
The specific guidance on immunogenicity assessment of biotherapeutic products has been elaborated by leading regulatory agencies such as the EMA and U.S. Food and Drug Administration (US FDA) [29, 30, 31, 32].
Validated analytical methods such as ELISA for quantification of biomarkers, drugs, biological products, and their metabolites in a given biological matrix (e.g. blood, plasma, serum, or urine) are critical for the successful conduct of nonclinical and clinical studies. Validating the analytical method ensures that the data are reliable [33]. Validated methods provide critical data to support the safety and effectiveness of drugs and biological products.
Although there is abundant literature relating to immunochemical methods, [34] EMEA [35, 36] and US FDA [8] have clearly defined the characteristics of the validation procedure for bioanalytical methods, which also applies to the validation of ELISAs, which are intended for use in diagnostics, toxicology, basic or applied research [37] or production control [38]. Metodology for the validation of bioanalytical methods must follow clear recomendations from reference institutions such as the EMEA [35, 39] or the WHO because that provides important measurements to be of satisfactory quality all over the world.
ELISA validation according to these recommendations means determining the following method caracteristics:
Specificity
Linearity –
Sensitivity
Accuracy
Precision (repetability = intra assay, inter assay, reproducibility = inter laboratory assay)
Robustnes
Acceptance criteria should be prospectively defined based on the intended use of the method.
Specificity means that the method must differentiate the targeted analyte from all other matrix components. Which is why it is important to test wether
Evaluation of specificity may be conducted during optimization and validation, when more data on the behavior of the analyte become available. Specificity should be tested with quality control (QC) samples [40]. QC samples are the samples with known amounts of the analyte, in identical matrix like the sample. These are usually in-house produced samples, with a lower amount of the analyte. When the method is performed with these QC samples and satisfactory results are obtained, then the method is also good, i.e. valid. If the method does not give good enough results with the QC samples, it means that the method is not of sufficient quality, so it must be investigated why the method worked poorly. The shortcomings must be corrected, and then again checked with QC samples. Still it needs to be defined what is satisfactory. The criterion for accepting the results obtained with QC samples is that the measured value does not deviate by more than 25% from the nominal value [40].
Linearity is the ability of the analytical method to produce results by calculating a direct proportion, within the working range. Linearity is described by range and detection limits.
Linearity is a function of values that can be graphically represented by a straight line. The linearity of an analytical method can be explained as its capability to show “results that are directly proportional to the concentration of the analyte in the sample” [39].
Unfortunately, the analytical response of a method is not always linear. Sometimes when the data are not linear they can be mathematically transformed, e.g. by applying logarithms but in some cases or some range of immunoassays transformation is not appropriate.
Linearity is important as it confirms the sensitivity of the method for the analysis of concentration within a defined range. According to the EMEA International Council for Harmonization ICH Q2(R1) guideline, linearity of a given response must be evaluated using a minimum of 5 concentrations of the analyte (multi-point calibration). Then, the collected data must be statistically analyzed, by performing regression analysis using the method of the least squares, in order to mathematically determine the line that best fits a set of data. For linearity, the results are required to be represented as linear equation (Eq. (1)).
In a linear regression line, the regression coefficient is the constant “k” that represents the rate of change of one variable “y” as a function of change in the other “x” (thus the slope), while “n” is the Y-intercept. The correlation coefficient r, a value without units, expresses the precision of the linearity fit of the experimental data. In case of a value being less than 0.95, it may either be a result of a broad spreading during measurement or due to a non-linear correlation. Often, the coefficient of determination (R2) is used, which is the square r. For most methods applied at R2 ≥ 0.98 can be achieved. If there is a perfect linear relationship, it has a value of 1 (100%). Linearity studies are important because they define the range of the method within which the results are obtained accurately and precisely.
To summarize, linearity is one major aspect in the quantitative method validation procedures. It describes the range of concentrations for which the method can function reliably. If the data are non-linear, transformation into a linear form may be performed, or the data can be accepted as is while demonstrating a clear relation between the analyte concentration and the measured absorbance [41].
Senzitivity or limit of detection, (LD) for ELISA is defined in the same way as for other bionalaytical methods. At this point, it is appropriate to underline the difference between the limit of detection (LD) and lower limit of quantification nominal (LLOQ). LD is the lowest analyte concentration that can be distinguished from the assay background, while the LLOQ is the lowest concentration at which the analyte can be quantitated at defined levels for precision and accuracy. LD is determined from standard deviation of the sample blank and the slope of the linear curve (Eq. (2)).
LD—LD (detection limit) nominal
k—slope of the linear curve Eq. (1)
SD(b)—standard deviation of the blank [39]
There are bioanalytical methods which have the same values for LD and LLOQ, but with ELISA, especially when biologycal samples are measure this is not the case, and LD is lower than LLOQ. For liretature reference of these terms one should read Armbruster and Pry [42].
The accuracy of an analytical method describes the closeness of the value determined by the method to the nominal concentration. In practice, as the reference material is precious and universally needed, the first step is to make a sufficient amount of the quality control (QC) samples, previously standardized against the reference material. Then the QC sample can be used for determining validation characteristics. Accuracy should be assessed on samples spiked with known amounts of the analyte, the QC samples. The accuracy can be expressed as the difference between the obtained experimnental value and the nominal value (which is acurate), using the absolute or even better the relative error.
Δxi—absolute error of individual measurement
μ— nominal value
xi—measured value
It is important to perform multiple measurements for a single sample, in order to present the absolute error as the mean value of absolute errors of individual measurements (Eq. (4)).
n—number of measurements
Δx—mean value of absolute or standard error
Because of the numerical nature, the absolute value of the difference does not give insight into its significance for the accuracy of measurement, so it is always important to calculate the relative error as well.
The level of accuracy must be determined for the whole range of the analytical procedure. Minimal requirements for this are three concentrations one close to ULOQ, one close to LLOQ and one in the middle of the range, each in triplicate.
Today it is common practise to develop an ELISA as an internal laboratory assay without the standards or the QC samples or for titration experiments for the determination of the last measurable dilution. In this situation there is no measurable quantitifier for accuracy testing. For accuracy to be calculated as % that shows how much the obtained results corresponds with the actual value, it is necessary to use concrete, absolute and measurable quantity such as analyte concentration. In practise this can be achieved [43] with inhibitory ELISA, which is based on the dependance of the absorbance on inhibitor concentration. The difference between the described calculations is in the reverse proportion, as described in the ELISA standardization section [37].
Precision is a validation characteristic which describes the reproducibility of the measurement, in other words the closeness of two measurements of the same sample. Precision is higher if the results are closer to one another. At first glance it is easy to confuse accuracy with precision, because in both cases it is about the absolute and the relative error of the obtained results. Figure 2 shows the difference between accuracy and precicion, where accuracy describes the deviation from the actual (nominal) value, while precision describes the deviation from the mean value. Precision is determined by simply repeating the measurement.
Accuracy and precision defined.
Standard deviation, relative standard deviation (coefficient of variation) and confidence interval should be reported for each type of precision (intra, intermediate or inter) investigated [35]. The three parameters are dependent on the closeness of individual results to the mean value, and give the complete picture of the precision of the test.
DEVIATION is the difference between the measured value from the mean value, and has the same units as the measured value (Eq. (6)).
x̅—mean value of repeated measurements of the same sample
xi—one measured value
di—deviation, the difference between the mean value and one measurement
Standard deviation is the mean value of all measurement deviations Eq. (7).
COEFFICIENT OF VARIATION CV (relative standard deviation) is standard deviation expressed in percentages and is calculated based on the measured mean value x̅ (Eq. (8)).
CONFIDENCE INTERVAL (CI) is the range of values within which the “actual” result is found. A CI of 95% means that if the measurement was to be repeated an infinite number of times, 95% of the results would fall within this range of values. For validation purpose, higher CI, 95% or 99% is needed, with optimal performance within the middle part of the range. A wide CI can be caused by small number of samples or by a large variance between sample measurements. Range of values for the given CI shows precision. This parameter is easily calculated by statistical programmes, or by a profesional statistician.
Intra-assay validation shows the reproducibility between wells within an assay plate. Data resulting from intra-assay validation helps ensure that repeated measurement of the same sample on a single plate gives comparable results. Repeatability should be assessed using a minimum of 6 determinations covering the specified range for the procedure (e.g. 3 concentrations, 2 replicates each), or a minimum of 6 determinations at 100% of the test concentration [39].
The % CV for each sample is calculated by finding the standard deviation of multiplicate results dividing that by the multiplicate mean, and multiplying the result by 100 (Eq. (8)). The average of the individual CVs is reported as the intra-assay CV (CVintra-assay).
Usually, CV intra-assay of 10% or less is considered satisfactory [44].
Intermediate precision (sometimes called within-lab reproducibility) shows the reproducibility between assays done on different days, or different plates. Satisfactory inter-assay precision is typically <10% [44].
For example, to monitor plate-to-plate variation the same samples are analyzed in quadruplicate on ten different plates. The plate means are calculated and then used to calculate the overall mean, standard deviation, and % CV. Overall % CV is calculated by dividing the SD of the plate means with mean of the plate means and multiplying by 100 (Eq. (8)). The average of the all plates % CV represents the inter-assay CV (CVintermediate). In order to monitor daily variation quadruplicate samples are analyzed in ten different days and analyzed in the same way.
Reproducibility is assessed by means of an inter-laboratory trial. The outcome of the cross validation is critical in determining whether the obtained data are reliable and whether they can be compared and used. Reproducibility should be considered in case of the standardization of an analytical procedure, for instance, for inclusion of procedures in pharmacopeias.
Satisfactory value for CVinter-assay is 10–15% [43].
Analyzing the literature it can be seen that the term inter assay is sometimes used for precision assessment on different days or on different plates, and sometimes for testing in different laboratories. Acording to EMEA, the term inter assay precision describes precision of the measurement assessment in different laboratories. If it is to be used in a different context it shold be described.
Robustness testing involves monitoring the effects of small unintentional errors on the quantitative and qualitative characteristics of the method, where the errors relate to the internal parameters described in the method prescription. For example, buffer temperature, incubation temperature, sample incubation time, secondary antibody incubation time, number of washes before color development, color development time, and the like. This feature shows the reliability of the method despite minor deviations in performance.
There is also the notion of rigidity - as a sub-notion of robustness - which monitors the effects of changes in external parameters such as other lots of chemicals, other people working, other instruments used and the like.
Practically, this property is not measured or calculated in a certain way, but is established during the development of the method (optimization). Data on this can also be collected during operation.
This guideline describes full validation methodology. In case when method is already validated, when a smaller change to the protocol is instated, a full validation may not be necessary. It is possible to perform partial validation, and the nature of the modification will determine the extent of validation required. All modifications should be reported and the scope of revalidation or partial validation justified [34].
In our experience ELISA is an excellent analytical method which can be used for the detection and quantification of numerous biomolecules. No matter what this specific biomolecule is, the basis of ELISA is the antigen–antibody interaction. The existence of this specific interaction usually enables the construction of different ELISA protocols, dependent on your prior knowledge and imagination. After careful protocol optimization, determination of validation characteristics and the acquirement of an appropriate standard you can get a reliable and inexpensive analytical method useful in diagnostics, research or biomedicine in general.
Work presented in this paper was supported by the Institute of Virology, Vaccines and Sera “Torlak” funds and by Ministry of Education, Science and Technological Development, Republic of Serbia.
The authors declare that they have no known competing financial interest or personal relationship that could have appeared to influence the work reported in this paper.
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Gharieb"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"10903",title:"Genetically Modified Plants and Beyond",subtitle:null,isOpenForSubmission:!1,hash:"4d7ed4faab99c92cd4d676dc86501df9",slug:"genetically-modified-plants-and-beyond",bookSignature:"Idah Sithole Niang",coverURL:"https://cdn.intechopen.com/books/images_new/10903.jpg",editedByType:"Edited by",publishedDate:"May 18th 2022",editors:[{id:"90172",title:"Prof.",name:"Idah",middleName:null,surname:"Sithole-Niang",slug:"idah-sithole-niang",fullName:"Idah Sithole-Niang"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"10904",title:"Fusarium",subtitle:"An Overview of the Genus",isOpenForSubmission:!1,hash:"49d9063e43f94bd1517d65fbc58b93c3",slug:"fusarium-an-overview-of-the-genus",bookSignature:"Seyed Mahyar Mirmajlessi",coverURL:"https://cdn.intechopen.com/books/images_new/10904.jpg",editedByType:"Edited by",publishedDate:"May 18th 2022",editors:[{id:"100573",title:"Dr.",name:"Seyed Mahyar",middleName:null,surname:"Mirmajlessi",slug:"seyed-mahyar-mirmajlessi",fullName:"Seyed Mahyar Mirmajlessi"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"10654",title:"Brain-Computer Interface",subtitle:null,isOpenForSubmission:!1,hash:"a5308884068cc53ed31c6baba756857f",slug:"brain-computer-interface",bookSignature:"Vahid Asadpour",coverURL:"https://cdn.intechopen.com/books/images_new/10654.jpg",editedByType:"Edited by",publishedDate:"May 18th 2022",editors:[{id:"165328",title:"Dr.",name:"Vahid",middleName:null,surname:"Asadpour",slug:"vahid-asadpour",fullName:"Vahid Asadpour"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"10676",title:"Recent Applications in Graph Theory",subtitle:null,isOpenForSubmission:!1,hash:"900c60742d224080732bd16bd25ccba8",slug:"recent-applications-in-graph-theory",bookSignature:"Harun Pirim",coverURL:"https://cdn.intechopen.com/books/images_new/10676.jpg",editedByType:"Edited by",publishedDate:"May 18th 2022",editors:[{id:"146092",title:"Dr.",name:"Harun",middleName:null,surname:"Pirim",slug:"harun-pirim",fullName:"Harun Pirim"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"11196",title:"New Updates in E-Learning",subtitle:null,isOpenForSubmission:!1,hash:"6afaadf68e2a0a4b370ac5ceb5ca89c6",slug:"new-updates-in-e-learning",bookSignature:"Eduard Babulak",coverURL:"https://cdn.intechopen.com/books/images_new/11196.jpg",editedByType:"Edited by",publishedDate:"May 18th 2022",editors:[{id:"10086",title:"Prof.",name:"Eduard",middleName:null,surname:"Babulak",slug:"eduard-babulak",fullName:"Eduard Babulak"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"9974",title:"E-Learning and Digital Education in the Twenty-First Century",subtitle:null,isOpenForSubmission:!1,hash:"88b58d66e975df20425fc1dfd22d53aa",slug:"e-learning-and-digital-education-in-the-twenty-first-century",bookSignature:"M. Mahruf C. Shohel",coverURL:"https://cdn.intechopen.com/books/images_new/9974.jpg",editedByType:"Edited by",publishedDate:"May 18th 2022",editors:[{id:"94099",title:"Dr.",name:"M. Mahruf C.",middleName:null,surname:"Shohel",slug:"m.-mahruf-c.-shohel",fullName:"M. Mahruf C. Shohel"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}}]},subject:{topic:{id:"424",title:"Phycology",slug:"phycology",parent:{id:"59",title:"Microbiology",slug:"biochemistry-genetics-and-molecular-biology-microbiology"},numberOfBooks:3,numberOfSeries:0,numberOfAuthorsAndEditors:56,numberOfWosCitations:100,numberOfCrossrefCitations:94,numberOfDimensionsCitations:258,videoUrl:null,fallbackUrl:null,description:null},booksByTopicFilter:{topicId:"424",sort:"-publishedDate",limit:12,offset:0},booksByTopicCollection:[{type:"book",id:"9354",title:"Microalgae",subtitle:"From Physiology to Application",isOpenForSubmission:!1,hash:"affa344272fbd8d5cd80cab53f814303",slug:"microalgae-from-physiology-to-application",bookSignature:"Milada Vítová",coverURL:"https://cdn.intechopen.com/books/images_new/9354.jpg",editedByType:"Edited by",editors:[{id:"253951",title:"Dr.",name:"Milada",middleName:null,surname:"Vítová",slug:"milada-vitova",fullName:"Milada Vítová"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"6889",title:"Algae",subtitle:null,isOpenForSubmission:!1,hash:"13797d037dd9bd0c3f7a232fff1c759d",slug:"algae",bookSignature:"Yee Keung Wong",coverURL:"https://cdn.intechopen.com/books/images_new/6889.jpg",editedByType:"Edited by",editors:[{id:"227706",title:"Dr.",name:"Yee Keung",middleName:null,surname:"Wong",slug:"yee-keung-wong",fullName:"Yee Keung Wong"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"5128",title:"Algae",subtitle:"Organisms for Imminent Biotechnology",isOpenForSubmission:!1,hash:"01a35d1259dcce526d25cf3f23237696",slug:"algae-organisms-for-imminent-biotechnology",bookSignature:"Nooruddin Thajuddin and Dharumadurai Dhanasekaran",coverURL:"https://cdn.intechopen.com/books/images_new/5128.jpg",editedByType:"Edited by",editors:[{id:"89852",title:"Dr.",name:"Nooruddin",middleName:null,surname:"Thajuddin",slug:"nooruddin-thajuddin",fullName:"Nooruddin Thajuddin"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}}],booksByTopicTotal:3,seriesByTopicCollection:[],seriesByTopicTotal:0,mostCitedChapters:[{id:"50544",doi:"10.5772/62909",title:"Algal Nanoparticles: Synthesis and Biotechnological Potentials",slug:"algal-nanoparticles-synthesis-and-biotechnological-potentials",totalDownloads:5784,totalCrossrefCites:17,totalDimensionsCites:67,abstract:"A nanoparticle can be defined as a small object that behaves as a whole unit in terms of its transport and properties. Nanoparticles are sized between 1 and 100 nm in diameter. Nanoparticles can act against the microbes in multiple ways, and the microbes are less likely to develop resistance against nanoparticles because it requires multiple gene mutations. The large surface-to-volume ratio of nanoparticles, their ability to easily interact with other particles, and several other features make them attractive tools in various fields. Nanoparticles are widely used various fields such as electronics, cosmetics, biomedical, and biotechnology. Nanoparticles can be synthesized by physical methods such as attrition, pyrolysis, and using some wet chemical methods. The physical and chemical methods have various drawbacks such as high cost of production, require high energy input and generation of toxic by-products. To overcome this, several biological methods are employed in the synthesis of nanoparticles. The biological methods are generally cost effective, nontoxic, and ecofriendly. This chapter focuses on the methods involved in algal-synthesized nanoparticles and its applications.",book:{id:"5128",slug:"algae-organisms-for-imminent-biotechnology",title:"Algae",fullTitle:"Algae - Organisms for Imminent Biotechnology"},signatures:"Felix LewisOscar, Sasikumar Vismaya, Manivel Arunkumar,\nNooruddin Thajuddin, Dharumadurai Dhanasekaran and Chari\nNithya",authors:[{id:"183668",title:"Dr.",name:"Nithya",middleName:null,surname:"Chari",slug:"nithya-chari",fullName:"Nithya Chari"}]},{id:"51074",doi:"10.5772/62916",title:"Algae as an Indicator of Water Quality",slug:"algae-as-an-indicator-of-water-quality",totalDownloads:5034,totalCrossrefCites:11,totalDimensionsCites:23,abstract:"The formation of plankton/algae under natural conditions is related to tolerance class (ecological optimum) due to abiotic limiting factors of ecosystem, as well as the biotic interactions among algae. In the ecological niche, the appearance of organisms is affected by anthropogenic and non-anthropogenic environmental factors. Algae composition and temporal variation in abundances are important in determining the trophic level of lakes. Algal communities are sensitive to changes in their habitat, and thus, total biomass of algae and many algae species are used as indicators of water quality. Algae communities give more knowledge on variations in water quality than nutrient or chlorophyll-a values. Water quality is a canonical group of physical, chemical, and biological properties of the given water. Consequently, eutrophication of freshwater is regarded as a water quality which results in the degeneration of the aquatic ecosystem and affects water utilisation. Cyanobacteria has been accepted as a major indicator of eutrophication in freshwater as their blooms are common in waters affected by nutrient concentration. The purpose of this chapter is to assess physical and chemical variables and the role of algal abundance to determine the water quality in the freshwater ecosystems.",book:{id:"5128",slug:"algae-organisms-for-imminent-biotechnology",title:"Algae",fullTitle:"Algae - Organisms for Imminent Biotechnology"},signatures:"Didem Gökçe",authors:[{id:"178260",title:"Associate Prof.",name:"Didem",middleName:null,surname:"Gokce",slug:"didem-gokce",fullName:"Didem Gokce"}]},{id:"50534",doi:"10.5772/63069",title:"Considerations for Photobioreactor Design and Operation for Mass Cultivation of Microalgae",slug:"considerations-for-photobioreactor-design-and-operation-for-mass-cultivation-of-microalgae",totalDownloads:5994,totalCrossrefCites:8,totalDimensionsCites:17,abstract:"Microalgae have great biotechnological potential for production of substances through photosynthesis. Light capture process and electron transportation imply energy losses due to reflection, fluorescence emission, and energy dissipation as heat, giving a maximum theoretical value of 8‐9% for microalgae energy capture efficiency and conversion to biomass. For development of full potential of microalgae the knowledge of the light capture process is required. High yields can only be obtained linking photobioreactor design with biological process taking place inside. In massive microalgae cultures, light gradients are generated and this depends on the biomass concentration, cellular types, cells sizes, and pigment content, and also on geometry, hydrodynamic, and light conditions inside the photobioreactor. In the present chapter we explain the relationship between light energy capture process and photobioreactor design and operation conditions, like turbulence, gas exchange, and nutrient requirements. Finally, the productivity and costs are discussed, and the parameters that determine the economic viability of any microalgae culture.",book:{id:"5128",slug:"algae-organisms-for-imminent-biotechnology",title:"Algae",fullTitle:"Algae - Organisms for Imminent Biotechnology"},signatures:"Juan Cristóbal García Cañedo and Gema Lorena López Lizárraga",authors:[{id:"185868",title:"Dr.",name:"Gema Lorena",middleName:null,surname:"López-Lizárraga",slug:"gema-lorena-lopez-lizarraga",fullName:"Gema Lorena López-Lizárraga"},{id:"293413",title:"Dr.",name:"Juan Cristóbal",middleName:null,surname:"García Cañedo",slug:"juan-cristobal-garcia-canedo",fullName:"Juan Cristóbal García Cañedo"}]},{id:"69201",doi:"10.5772/intechopen.89324",title:"Drying and Quality of Microalgal Powders for Human Alimentation",slug:"drying-and-quality-of-microalgal-powders-for-human-alimentation",totalDownloads:1263,totalCrossrefCites:5,totalDimensionsCites:15,abstract:"The demand for natural foods with high protein content and functional properties is constantly growing in the last years. In this context, microalgae as Spirulina (Arthrospira spp.), Chlorella spp., Haematococcus pluvialis, Dunaliella salina, and others, assume a key role to diversify the offer of nutritious and functional ingredients and supplements. Microalgae are commercialized, mostly, as dried powders to facilitate their use as food ingredients and to allow easy transportation and long-term stability. Microalgal powder quality and storage stability depend mainly on drying method, packaging, and storage conditions. Most of the studies that approach the subject of microalgal drying evaluate the efficiency of the process and suitability for this raw material. However, studies that assess the effect of traditional and innovative drying methods on quality of microalgal powder for human consumption are rare in literature. In this chapter, the state of the art of drying processing technology for microalgae was reviewed, discussing the effect of dehydration on quality and stability of microalgal powders with potential use in human alimentation.",book:{id:"9354",slug:"microalgae-from-physiology-to-application",title:"Microalgae",fullTitle:"Microalgae - From Physiology to Application"},signatures:"Fábio de Farias Neves, Mariana Demarco and Giustino Tribuzi",authors:null},{id:"50671",doi:"10.5772/63272",title:"Challenges and Opportunities in the Present Era of Marine Algal Applications",slug:"challenges-and-opportunities-in-the-present-era-of-marine-algal-applications",totalDownloads:2747,totalCrossrefCites:3,totalDimensionsCites:12,abstract:"Marine algae are of high importance in their natural habitats and even more now in the world of green technology. The sprouting interest of the scientific community and industries in these organisms is driven by the fast-growing world of modern biotechnology. Genomics, transcriptomics, proteomics, metabolomics and their integration collectively termed here as ‘marine algal-omics’ have broadened the research horizon in view of enhancing human’s life by addressing environmental problems and encouraging novelty in the field of pharmaceuticals among so many more. Their use in the human society dates back to 500 B. C. in China and later across the globe; they are still being used for similar purposes and more today. There is a hiking interest in marine algae and their derivatives—from phycoremediation, food supplements, pharmaceuticals to dyes. Marine algae are currently considered as an emerging panacea for the society. They are being studied in a multitude of arenas. The multi-use of marine algae is enticing and promises to be a boon for industrial applications. Yet, most marine algae face challenges that might variably constrain their commercialisation. This chapter gives an overview of marine algae including all the ‘omics’ technologies involved in studying marine algae and it explores their multitude applications. It also draws the various successful industries budded around them and presents some of the challenges and opportunities along with future directions.",book:{id:"5128",slug:"algae-organisms-for-imminent-biotechnology",title:"Algae",fullTitle:"Algae - Organisms for Imminent Biotechnology"},signatures:"Keshini Beetul, Arvind Gopeechund, Deepeeka Kaullysing, Sushma\nMattan-Moorgawa, Daneshwar Puchooa and Ranjeet Bhagooli",authors:[{id:"178209",title:"Ms.",name:"Keshini",middleName:null,surname:"Beetul",slug:"keshini-beetul",fullName:"Keshini Beetul"},{id:"184390",title:"Mr.",name:"Arvind",middleName:null,surname:"Gopeechund",slug:"arvind-gopeechund",fullName:"Arvind Gopeechund"},{id:"184391",title:"Ms.",name:"Deepeeka",middleName:null,surname:"Kaullysing",slug:"deepeeka-kaullysing",fullName:"Deepeeka Kaullysing"},{id:"184392",title:"Mrs.",name:"Sushma",middleName:null,surname:"Mattan-Moorgawa",slug:"sushma-mattan-moorgawa",fullName:"Sushma Mattan-Moorgawa"},{id:"184393",title:"Prof.",name:"Daneshwar",middleName:null,surname:"Puchooa",slug:"daneshwar-puchooa",fullName:"Daneshwar Puchooa"},{id:"184394",title:"Dr.",name:"Ranjeet",middleName:null,surname:"Bhagooli",slug:"ranjeet-bhagooli",fullName:"Ranjeet Bhagooli"}]}],mostDownloadedChaptersLast30Days:[{id:"64156",title:"Cyanobacteria Growth Kinetics",slug:"cyanobacteria-growth-kinetics",totalDownloads:1799,totalCrossrefCites:2,totalDimensionsCites:5,abstract:"Harmful cyanobacterial blooms are a global problem for freshwater ecosystems used for drinking water supply and recreational purposes. Cyanobacteria also produce a wide variety of toxic secondary metabolites, called cyanotoxins. High water temperatures have been known to lead to cyanobacterial bloom development in temperate and semiarid regions. Increased temperatures as a result of climate change could therefore favor the growth of cyanobacteria, thus augmenting the risks associated with the blooms. Though temperature is the main factor affecting the growth kinetics of bacteria, the availability of nutrients such as nitrogen and phosphorus also plays a significant role. This chapter studies the growth kinetics of toxin-producing Microcystis aeruginosa and evaluates potential risks to the population in scenarios of climate change and the presence of nutrients. The most suitable control methods for mitigation are also evaluated.",book:{id:"6889",slug:"algae",title:"Algae",fullTitle:"Algae"},signatures:"Leda Giannuzzi",authors:[{id:"252117",title:"Dr.",name:"Leda",middleName:null,surname:"Giannuzzi",slug:"leda-giannuzzi",fullName:"Leda Giannuzzi"}]},{id:"65952",title:"CO2 Capture for Industries by Algae",slug:"co-sub-2-sub-capture-for-industries-by-algae",totalDownloads:2102,totalCrossrefCites:5,totalDimensionsCites:11,abstract:"The increased usage of fossil fuels has led to increase in the concentration of CO2, which is a greenhouse gas responsible for global warming. Algae-based CO2 conversion is a cost-effective option for reducing carbon footprint. In addition, algae-based CO2 mitigation strategy has the potential to obtain valuable products at the end of the process. In the present study, freshwater algal species were isolated and identified for CO2 capture, such as Hydrodictyon, Spirogyra, Oscillatoria, Oedogonium, and Chlorella. The algal strains were screened based on different parameters like fast growth rate, high rate of photosynthesis, strong tolerance to the trace constituents of other gases (gaseous hydrocarbons, NOx, SOx, etc.), high temperature tolerance, and possibility to produce high value products, etc. The study involves integrated methods for utilizing 90–99% CO2 from a natural gas processing industry (GAIL India, Ltd.) as well as 13–15% of CO2 from flue gas of thermal power plants (Chandrapura and Santaldih Thermal Power Station) as carbon nutrient source along with the additional nutritional supplements. A 400-ml and 25-l flat panel photo-bioreactor (PSI Photo-bioreactors) was used for CO2 capture. After CO2 capture, the algal biomass was used to extract value-added products such as amino acid rich feed, algal oil, algal pellets, etc.",book:{id:"6889",slug:"algae",title:"Algae",fullTitle:"Algae"},signatures:"Vetrivel Anguselvi, Reginald Ebhin Masto, Ashis Mukherjee and Pradeep Kumar Singh",authors:[{id:"255851",title:"Dr.",name:"Vetrivel",middleName:null,surname:"Anguselvi",slug:"vetrivel-anguselvi",fullName:"Vetrivel Anguselvi"},{id:"269996",title:"Dr.",name:"R E",middleName:null,surname:"Masto",slug:"r-e-masto",fullName:"R E Masto"},{id:"269997",title:"Dr.",name:"Ashis",middleName:null,surname:"Mukherjee",slug:"ashis-mukherjee",fullName:"Ashis Mukherjee"},{id:"270059",title:"Dr.",name:"P K",middleName:null,surname:"Singh",slug:"p-k-singh",fullName:"P K Singh"}]},{id:"51074",title:"Algae as an Indicator of Water Quality",slug:"algae-as-an-indicator-of-water-quality",totalDownloads:5034,totalCrossrefCites:11,totalDimensionsCites:23,abstract:"The formation of plankton/algae under natural conditions is related to tolerance class (ecological optimum) due to abiotic limiting factors of ecosystem, as well as the biotic interactions among algae. In the ecological niche, the appearance of organisms is affected by anthropogenic and non-anthropogenic environmental factors. Algae composition and temporal variation in abundances are important in determining the trophic level of lakes. Algal communities are sensitive to changes in their habitat, and thus, total biomass of algae and many algae species are used as indicators of water quality. Algae communities give more knowledge on variations in water quality than nutrient or chlorophyll-a values. Water quality is a canonical group of physical, chemical, and biological properties of the given water. Consequently, eutrophication of freshwater is regarded as a water quality which results in the degeneration of the aquatic ecosystem and affects water utilisation. Cyanobacteria has been accepted as a major indicator of eutrophication in freshwater as their blooms are common in waters affected by nutrient concentration. The purpose of this chapter is to assess physical and chemical variables and the role of algal abundance to determine the water quality in the freshwater ecosystems.",book:{id:"5128",slug:"algae-organisms-for-imminent-biotechnology",title:"Algae",fullTitle:"Algae - Organisms for Imminent Biotechnology"},signatures:"Didem Gökçe",authors:[{id:"178260",title:"Associate Prof.",name:"Didem",middleName:null,surname:"Gokce",slug:"didem-gokce",fullName:"Didem Gokce"}]},{id:"64455",title:"Cyanobacteria for PHB Bioplastics Production: A Review",slug:"cyanobacteria-for-phb-bioplastics-production-a-review",totalDownloads:2186,totalCrossrefCites:4,totalDimensionsCites:11,abstract:"Cyanobacteria, or blue-green algae, can be used as host to produce polyhydroxyalkanoates (PHA), which are promising bioplastic raw materials. The most important material thereof is polyhydroxybutyrate (PHB), which can replace the commodity polymer polypropylene (PP) in many applications, yielding a bio-based, biodegradable alternative solution. The advantage from using cyanobacteria to make PHB over the standard fermentation processes, with sugar or other organic (waste) materials as feedstock, is that the sustainability is better (compare first-generation biofuels with the feed vs. fuel debate), with CO2 being the only carbon source and sunlight being the sole energy source. In this review article, the state of the art of cyanobacterial PHB production and its outlook is discussed. Thirty-seven percent of dry cell weight of PHB could be obtained in 2018, which is getting close to up to 78% of PHB dry cell weight in heterotrophic microorganisms in fermentation reactors. A good potential for cyanobacterial PHB is seen throughout the literature.",book:{id:"6889",slug:"algae",title:"Algae",fullTitle:"Algae"},signatures:"Erich Markl, Hannes Grünbichler and Maximilian Lackner",authors:[{id:"251081",title:"Dr.",name:"Maximilian",middleName:null,surname:"Lackner",slug:"maximilian-lackner",fullName:"Maximilian Lackner"},{id:"255232",title:"Prof.",name:"Erich",middleName:null,surname:"Markl",slug:"erich-markl",fullName:"Erich Markl"},{id:"277237",title:"Dr.",name:"Hannes",middleName:null,surname:"Grünbichler",slug:"hannes-grunbichler",fullName:"Hannes Grünbichler"}]},{id:"50544",title:"Algal Nanoparticles: Synthesis and Biotechnological Potentials",slug:"algal-nanoparticles-synthesis-and-biotechnological-potentials",totalDownloads:5784,totalCrossrefCites:17,totalDimensionsCites:67,abstract:"A nanoparticle can be defined as a small object that behaves as a whole unit in terms of its transport and properties. Nanoparticles are sized between 1 and 100 nm in diameter. Nanoparticles can act against the microbes in multiple ways, and the microbes are less likely to develop resistance against nanoparticles because it requires multiple gene mutations. The large surface-to-volume ratio of nanoparticles, their ability to easily interact with other particles, and several other features make them attractive tools in various fields. Nanoparticles are widely used various fields such as electronics, cosmetics, biomedical, and biotechnology. Nanoparticles can be synthesized by physical methods such as attrition, pyrolysis, and using some wet chemical methods. The physical and chemical methods have various drawbacks such as high cost of production, require high energy input and generation of toxic by-products. To overcome this, several biological methods are employed in the synthesis of nanoparticles. The biological methods are generally cost effective, nontoxic, and ecofriendly. 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Saxena is a vice dean and professor at King George's Medical University, Lucknow, India. His research interests involve understanding the molecular mechanisms of host defense during human viral infections and developing new predictive, preventive, and therapeutic strategies for them using Japanese encephalitis virus (JEV), HIV, and emerging viruses as a model via stem cell and cell culture technologies. His research work has been published in various high-impact factor journals (Science, PNAS, Nature Medicine) with a high number of citations. He has received many awards and honors in India and abroad including various Young Scientist Awards, BBSRC India Partnering Award, and Dr. JC Bose National Award of Department of Biotechnology, Min. of Science and Technology, Govt. of India. 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Patil Medical College and Director, Centre for Advanced Medical Research (CAMR), BLDE (Deemed to be University), Vijayapur, Karnataka, India. Dr. Das did his M.S. and Ph.D. in Human Physiology from the University of Calcutta, Kolkata. His area of research is focused on understanding of molecular mechanisms of heavy metal activated low oxygen sensing pathways in vascular pathophysiology. He has invented a new method of estimation of serum vitamin E. His expertise in critical experimental protocols on vascular functions in experimental animals was well documented by his quality of publications. He was a Visiting Professor of Medicine at University of Leeds, United Kingdom (2014-2016) and Tulane University, New Orleans, USA (2017). For his immense contribution in medical research Ministry of Science and Technology, Government of India conferred him 'G.P. Chatterjee Memorial Research Prize-2019” and he is also the recipient of 'Dr.Raja Ramanna State Scientist Award 2015” by Government of Karnataka. He is a Fellow of the Royal Society of Biology (FRSB), London and Honorary Fellow of Karnataka Science and Technology Academy, Department of Science and Technology, Government of Karnataka.",institutionString:"BLDE (Deemed to be University), India",institution:null},{id:"243660",title:"Dr.",name:"Mallanagouda Shivanagouda",middleName:null,surname:"Biradar",slug:"mallanagouda-shivanagouda-biradar",fullName:"Mallanagouda Shivanagouda Biradar",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/243660/images/system/243660.jpeg",biography:"M. S. Biradar is Vice Chancellor and Professor of Medicine of\nBLDE (Deemed to be University), Vijayapura, Karnataka, India.\nHe obtained his MD with a gold medal in General Medicine and\nhas devoted himself to medical teaching, research, and administrations. He has also immensely contributed to medical research\non vascular medicine, which is reflected by his numerous publications including books and book chapters. Professor Biradar was\nalso Visiting Professor at Tulane University School of Medicine, New Orleans, USA.",institutionString:"BLDE (Deemed to be University)",institution:{name:"BLDE University",country:{name:"India"}}},{id:"289796",title:"Dr.",name:"Swastika",middleName:null,surname:"Das",slug:"swastika-das",fullName:"Swastika Das",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/289796/images/system/289796.jpeg",biography:"Swastika N. Das is Professor of Chemistry at the V. P. Dr. P. G.\nHalakatti College of Engineering and Technology, BLDE (Deemed\nto be University), Vijayapura, Karnataka, India. She obtained an\nMSc, MPhil, and PhD in Chemistry from Sambalpur University,\nOdisha, India. Her areas of research interest are medicinal chemistry, chemical kinetics, and free radical chemistry. She is a member\nof the investigators who invented a new modified method of estimation of serum vitamin E. She has authored numerous publications including book\nchapters and is a mentor of doctoral curriculum at her university.",institutionString:"BLDEA’s V.P.Dr.P.G.Halakatti College of Engineering & Technology",institution:{name:"BLDE University",country:{name:"India"}}},{id:"248459",title:"Dr.",name:"Akikazu",middleName:null,surname:"Takada",slug:"akikazu-takada",fullName:"Akikazu Takada",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/248459/images/system/248459.png",biography:"Akikazu Takada was born in Japan, 1935. After graduation from\nKeio University School of Medicine and finishing his post-graduate studies, he worked at Roswell Park Memorial Institute NY,\nUSA. He then took a professorship at Hamamatsu University\nSchool of Medicine. In thrombosis studies, he found the SK\npotentiator that enhances plasminogen activation by streptokinase. He is very much interested in simultaneous measurements\nof fatty acids, amino acids, and tryptophan degradation products. By using fatty\nacid analyses, he indicated that plasma levels of trans-fatty acids of old men were\nfar higher in the US than Japanese men. . He also showed that eicosapentaenoic acid\n(EPA) and docosahexaenoic acid (DHA) levels are higher, and arachidonic acid\nlevels are lower in Japanese than US people. By using simultaneous LC/MS analyses\nof plasma levels of tryptophan metabolites, he recently found that plasma levels of\nserotonin, kynurenine, or 5-HIAA were higher in patients of mono- and bipolar\ndepression, which are significantly different from observations reported before. In\nview of recent reports that plasma tryptophan metabolites are mainly produced by\nmicrobiota. He is now working on the relationships between microbiota and depression or autism.",institutionString:"Hamamatsu University School of Medicine",institution:{name:"Hamamatsu University School of Medicine",country:{name:"Japan"}}},{id:"137240",title:"Prof.",name:"Mohammed",middleName:null,surname:"Khalid",slug:"mohammed-khalid",fullName:"Mohammed Khalid",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/137240/images/system/137240.png",biography:"Mohammed Khalid received his B.S. degree in chemistry in 2000 and Ph.D. degree in physical chemistry in 2007 from the University of Khartoum, Sudan. He moved to School of Chemistry, Faculty of Science, University of Sydney, Australia in 2009 and joined Dr. Ron Clarke as a postdoctoral fellow where he worked on the interaction of ATP with the phosphoenzyme of the Na+/K+-ATPase and dual mechanisms of allosteric acceleration of the Na+/K+-ATPase by ATP; then he went back to Department of Chemistry, University of Khartoum as an assistant professor, and in 2014 he was promoted as an associate professor. In 2011, he joined the staff of Department of Chemistry at Taif University, Saudi Arabia, where he is currently an assistant professor. His research interests include the following: P-Type ATPase enzyme kinetics and mechanisms, kinetics and mechanisms of redox reactions, autocatalytic reactions, computational enzyme kinetics, allosteric acceleration of P-type ATPases by ATP, exploring of allosteric sites of ATPases, and interaction of ATP with ATPases located in cell membranes.",institutionString:"Taif University",institution:{name:"Taif University",country:{name:"Saudi Arabia"}}},{id:"63810",title:"Prof.",name:"Jorge",middleName:null,surname:"Morales-Montor",slug:"jorge-morales-montor",fullName:"Jorge Morales-Montor",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/63810/images/system/63810.png",biography:"Dr. Jorge Morales-Montor was recognized with the Lola and Igo Flisser PUIS Award for best graduate thesis at the national level in the field of parasitology. He received a fellowship from the Fogarty Foundation to perform postdoctoral research stay at the University of Georgia. He has 153 journal articles to his credit. He has also edited several books and published more than fifty-five book chapters. He is a member of the Mexican Academy of Sciences, Latin American Academy of Sciences, and the National Academy of Medicine. He has received more than thirty-five awards and has supervised numerous bachelor’s, master’s, and Ph.D. students. Dr. Morales-Montor is the past president of the Mexican Society of Parasitology.",institutionString:"National Autonomous University of Mexico",institution:{name:"National Autonomous University of Mexico",country:{name:"Mexico"}}},{id:"217215",title:"Dr.",name:"Palash",middleName:null,surname:"Mandal",slug:"palash-mandal",fullName:"Palash Mandal",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/217215/images/system/217215.jpeg",biography:null,institutionString:"Charusat University",institution:null},{id:"49739",title:"Dr.",name:"Leszek",middleName:null,surname:"Szablewski",slug:"leszek-szablewski",fullName:"Leszek Szablewski",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/49739/images/system/49739.jpg",biography:"Leszek Szablewski is a professor of medical sciences. He received his M.S. in the Faculty of Biology from the University of Warsaw and his PhD degree from the Institute of Experimental Biology Polish Academy of Sciences. He habilitated in the Medical University of Warsaw, and he obtained his degree of Professor from the President of Poland. Professor Szablewski is the Head of Chair and Department of General Biology and Parasitology, Medical University of Warsaw. Professor Szablewski has published over 80 peer-reviewed papers in journals such as Journal of Alzheimer’s Disease, Biochim. Biophys. Acta Reviews of Cancer, Biol. Chem., J. Biomed. Sci., and Diabetes/Metabol. Res. Rev, Endocrine. He is the author of two books and four book chapters. He has edited four books, written 15 scripts for students, is the ad hoc reviewer of over 30 peer-reviewed journals, and editorial member of peer-reviewed journals. Prof. Szablewski’s research focuses on cell physiology, genetics, and pathophysiology. He works on the damage caused by lack of glucose homeostasis and changes in the expression and/or function of glucose transporters due to various diseases. He has given lectures, seminars, and exercises for students at the Medical University.",institutionString:"Medical University of Warsaw",institution:{name:"Medical University of Warsaw",country:{name:"Poland"}}},{id:"173123",title:"Dr.",name:"Maitham",middleName:null,surname:"Khajah",slug:"maitham-khajah",fullName:"Maitham Khajah",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/173123/images/system/173123.jpeg",biography:"Dr. Maitham A. Khajah received his degree in Pharmacy from Faculty of Pharmacy, Kuwait University, in 2003 and obtained his PhD degree in December 2009 from the University of Calgary, Canada (Gastrointestinal Science and Immunology). Since January 2010 he has been assistant professor in Kuwait University, Faculty of Pharmacy, Department of Pharmacology and Therapeutics. His research interest are molecular targets for the treatment of inflammatory bowel disease (IBD) and the mechanisms responsible for immune cell chemotaxis. He cosupervised many students for the MSc Molecular Biology Program, College of Graduate Studies, Kuwait University. Ever since joining Kuwait University in 2010, he got various grants as PI and Co-I. He was awarded the Best Young Researcher Award by Kuwait University, Research Sector, for the Year 2013–2014. He was a member in the organizing committee for three conferences organized by Kuwait University, Faculty of Pharmacy, as cochair and a member in the scientific committee (the 3rd, 4th, and 5th Kuwait International Pharmacy Conference).",institutionString:"Kuwait University",institution:{name:"Kuwait University",country:{name:"Kuwait"}}},{id:"195136",title:"Dr.",name:"Aya",middleName:null,surname:"Adel",slug:"aya-adel",fullName:"Aya Adel",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/195136/images/system/195136.jpg",biography:"Dr. Adel works as an Assistant Lecturer in the unit of Phoniatrics, Department of Otolaryngology, Ain Shams University in Cairo, Egypt. Dr. Adel is especially interested in joint attention and its impairment in autism spectrum disorder",institutionString:"Ain Shams University",institution:{name:"Ain Shams University",country:{name:"Egypt"}}},{id:"94911",title:"Dr.",name:"Boulenouar",middleName:null,surname:"Mesraoua",slug:"boulenouar-mesraoua",fullName:"Boulenouar Mesraoua",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/94911/images/system/94911.png",biography:"Dr Boulenouar Mesraoua is the Associate Professor of Clinical Neurology at Weill Cornell Medical College-Qatar and a Consultant Neurologist at Hamad Medical Corporation at the Neuroscience Department; He graduated as a Medical Doctor from the University of Oran, Algeria; he then moved to Belgium, the City of Liege, for a Residency in Internal Medicine and Neurology at Liege University; after getting the Belgian Board of Neurology (with high marks), he went to the National Hospital for Nervous Diseases, Queen Square, London, United Kingdom for a fellowship in Clinical Neurophysiology, under Pr Willison ; Dr Mesraoua had also further training in Epilepsy and Continuous EEG Monitoring for two years (from 2001-2003) in the Neurophysiology department of Zurich University, Switzerland, under late Pr Hans Gregor Wieser ,an internationally known epileptologist expert. \n\nDr B. Mesraoua is the Director of the Neurology Fellowship Program at the Neurology Section and an active member of the newly created Comprehensive Epilepsy Program at Hamad General Hospital, Doha, Qatar; he is also Assistant Director of the Residency Program at the Qatar Medical School. \nDr B. Mesraoua's main interests are Epilepsy, Multiple Sclerosis, and Clinical Neurology; He is the Chairman and the Organizer of the well known Qatar Epilepsy Symposium, he is running yearly for the past 14 years and which is considered a landmark in the Gulf region; He has also started last year , together with other epileptologists from Qatar, the region and elsewhere, a yearly International Epilepsy School Course, which was attended by many neurologists from the Area.\n\nInternationally, Dr Mesraoua is an active and elected member of the Commission on Eastern Mediterranean Region (EMR ) , a regional branch of the International League Against Epilepsy (ILAE), where he represents the Middle East and North Africa(MENA ) and where he holds the position of chief of the Epilepsy Epidemiology Section; Dr Mesraoua is a member of the American Academy of Neurology, the Europeen Academy of Neurology and the American Epilepsy Society.\n\nDr Mesraoua's main objectives are to encourage frequent gathering of the epileptologists/neurologists from the MENA region and the rest of the world, promote Epilepsy Teaching in the MENA Region, and encourage multicenter studies involving neurologists and epileptologists in the MENA region, particularly epilepsy epidemiological studies. \n\nDr. Mesraoua is the recipient of two research Grants, as the Lead Principal Investigator (750.000 USD and 250.000 USD) from the Qatar National Research Fund (QNRF) and the Hamad Hospital Internal Research Grant (IRGC), on the following topics : “Continuous EEG Monitoring in the ICU “ and on “Alpha-lactoalbumin , proof of concept in the treatment of epilepsy” .Dr Mesraoua is a reviewer for the journal \"seizures\" (Europeen Epilepsy Journal ) as well as dove journals ; Dr Mesraoua is the author and co-author of many peer reviewed publications and four book chapters in the field of Epilepsy and Clinical Neurology",institutionString:"Weill Cornell Medical College in Qatar",institution:{name:"Weill Cornell Medical College in Qatar",country:{name:"Qatar"}}},{id:"282429",title:"Prof.",name:"Covanis",middleName:null,surname:"Athanasios",slug:"covanis-athanasios",fullName:"Covanis Athanasios",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/282429/images/system/282429.jpg",biography:null,institutionString:"Neurology-Neurophysiology Department of the Children Hospital Agia Sophia",institution:null},{id:"190980",title:"Prof.",name:"Marwa",middleName:null,surname:"Mahmoud Saleh",slug:"marwa-mahmoud-saleh",fullName:"Marwa Mahmoud Saleh",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/190980/images/system/190980.jpg",biography:"Professor Marwa Mahmoud Saleh is a doctor of medicine and currently works in the unit of Phoniatrics, Department of Otolaryngology, Ain Shams University in Cairo, Egypt. She got her doctoral degree in 1991 and her doctoral thesis was accomplished in the University of Iowa, United States. Her publications covered a multitude of topics as videokymography, cochlear implants, stuttering, and dysphagia. She has lectured Egyptian phonology for many years. Her recent research interest is joint attention in autism.",institutionString:"Ain Shams University",institution:{name:"Ain Shams University",country:{name:"Egypt"}}},{id:"259190",title:"Dr.",name:"Syed Ali Raza",middleName:null,surname:"Naqvi",slug:"syed-ali-raza-naqvi",fullName:"Syed Ali Raza Naqvi",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/259190/images/system/259190.png",biography:"Dr. Naqvi is a radioanalytical chemist and is working as an associate professor of analytical chemistry in the Department of Chemistry, Government College University, Faisalabad, Pakistan. Advance separation techniques, nuclear analytical techniques and radiopharmaceutical analysis are the main courses that he is teaching to graduate and post-graduate students. In the research area, he is focusing on the development of organic- and biomolecule-based radiopharmaceuticals for diagnosis and therapy of infectious and cancerous diseases. Under the supervision of Dr. Naqvi, three students have completed their Ph.D. degrees and 41 students have completed their MS degrees. He has completed three research projects and is currently working on 2 projects entitled “Radiolabeling of fluoroquinolone derivatives for the diagnosis of deep-seated bacterial infections” and “Radiolabeled minigastrin peptides for diagnosis and therapy of NETs”. He has published about 100 research articles in international reputed journals and 7 book chapters. Pakistan Institute of Nuclear Science & Technology (PINSTECH) Islamabad, Punjab Institute of Nuclear Medicine (PINM), Faisalabad and Institute of Nuclear Medicine and Radiology (INOR) Abbottabad are the main collaborating institutes.",institutionString:"Government College University",institution:{name:"Government College University, Faisalabad",country:{name:"Pakistan"}}},{id:"58390",title:"Dr.",name:"Gyula",middleName:null,surname:"Mozsik",slug:"gyula-mozsik",fullName:"Gyula Mozsik",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/58390/images/system/58390.png",biography:"Gyula Mózsik MD, Ph.D., ScD (med), is an emeritus professor of Medicine at the First Department of Medicine, Univesity of Pécs, Hungary. He was head of this department from 1993 to 2003. His specializations are medicine, gastroenterology, clinical pharmacology, clinical nutrition, and dietetics. His research fields are biochemical pharmacological examinations in the human gastrointestinal (GI) mucosa, mechanisms of retinoids, drugs, capsaicin-sensitive afferent nerves, and innovative pharmacological, pharmaceutical, and nutritional (dietary) research in humans. He has published about 360 peer-reviewed papers, 197 book chapters, 692 abstracts, 19 monographs, and has edited 37 books. He has given about 1120 regular and review lectures. He has organized thirty-eight national and international congresses and symposia. He is the founder of the International Conference on Ulcer Research (ICUR); International Union of Pharmacology, Gastrointestinal Section (IUPHAR-GI); Brain-Gut Society symposiums, and gastrointestinal cytoprotective symposiums. He received the Andre Robert Award from IUPHAR-GI in 2014. Fifteen of his students have been appointed as full professors in Egypt, Cuba, and Hungary.",institutionString:"University of Pécs",institution:{name:"University of Pecs",country:{name:"Hungary"}}},{id:"277367",title:"M.Sc.",name:"Daniel",middleName:"Martin",surname:"Márquez López",slug:"daniel-marquez-lopez",fullName:"Daniel Márquez López",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/277367/images/7909_n.jpg",biography:"Msc Daniel Martin Márquez López has a bachelor degree in Industrial Chemical Engineering, a Master of science degree in the same área and he is a PhD candidate for the Instituto Politécnico Nacional. His Works are realted to the Green chemistry field, biolubricants, biodiesel, transesterification reactions for biodiesel production and the manipulation of oils for therapeutic purposes.",institutionString:null,institution:{name:"Instituto Politécnico Nacional",country:{name:"Mexico"}}},{id:"196544",title:"Prof.",name:"Angel",middleName:null,surname:"Catala",slug:"angel-catala",fullName:"Angel Catala",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/196544/images/system/196544.jpg",biography:"Angel Catalá studied chemistry at Universidad Nacional de La Plata, Argentina, where he received a Ph.D. in Chemistry (Biological Branch) in 1965. From 1964 to 1974, he worked as an Assistant in Biochemistry at the School of Medicine at the same university. From 1974 to 1976, he was a fellow of the National Institutes of Health (NIH) at the University of Connecticut, Health Center, USA. From 1985 to 2004, he served as a Full Professor of Biochemistry at the Universidad Nacional de La Plata. He is a member of the National Research Council (CONICET), Argentina, and the Argentine Society for Biochemistry and Molecular Biology (SAIB). His laboratory has been interested for many years in the lipid peroxidation of biological membranes from various tissues and different species. Dr. Catalá has directed twelve doctoral theses, published more than 100 papers in peer-reviewed journals, several chapters in books, and edited twelve books. He received awards at the 40th International Conference Biochemistry of Lipids 1999 in Dijon, France. He is the winner of the Bimbo Pan-American Nutrition, Food Science and Technology Award 2006 and 2012, South America, Human Nutrition, Professional Category. In 2006, he won the Bernardo Houssay award in pharmacology, in recognition of his meritorious works of research. Dr. Catalá belongs to the editorial board of several journals including Journal of Lipids; International Review of Biophysical Chemistry; Frontiers in Membrane Physiology and Biophysics; World Journal of Experimental Medicine and Biochemistry Research International; World Journal of Biological Chemistry, Diabetes, and the Pancreas; International Journal of Chronic Diseases & Therapy; and International Journal of Nutrition. He is the co-editor of The Open Biology Journal and associate editor for Oxidative Medicine and Cellular Longevity.",institutionString:"Universidad Nacional de La Plata",institution:{name:"National University of La Plata",country:{name:"Argentina"}}},{id:"186585",title:"Dr.",name:"Francisco Javier",middleName:null,surname:"Martin-Romero",slug:"francisco-javier-martin-romero",fullName:"Francisco Javier Martin-Romero",position:null,profilePictureURL:"https://s3.us-east-1.amazonaws.com/intech-files/0030O00002bSB3HQAW/Profile_Picture_1631258137641",biography:"Francisco Javier Martín-Romero (Javier) is a Professor of Biochemistry and Molecular Biology at the University of Extremadura, Spain. He is also a group leader at the Biomarkers Institute of Molecular Pathology. Javier received his Ph.D. in 1998 in Biochemistry and Biophysics. At the National Cancer Institute (National Institute of Health, Bethesda, MD) he worked as a research associate on the molecular biology of selenium and its role in health and disease. After postdoctoral collaborations with Carlos Gutierrez-Merino (University of Extremadura, Spain) and Dario Alessi (University of Dundee, UK), he established his own laboratory in 2008. The interest of Javier's lab is the study of cell signaling with a special focus on Ca2+ signaling, and how Ca2+ transport modulates the cytoskeleton, migration, differentiation, cell death, etc. He is especially interested in the study of Ca2+ channels, and the role of STIM1 in the initiation of pathological events.",institutionString:null,institution:{name:"University of Extremadura",country:{name:"Spain"}}},{id:"217323",title:"Prof.",name:"Guang-Jer",middleName:null,surname:"Wu",slug:"guang-jer-wu",fullName:"Guang-Jer Wu",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/217323/images/8027_n.jpg",biography:null,institutionString:null,institution:null},{id:"148546",title:"Dr.",name:"Norma Francenia",middleName:null,surname:"Santos-Sánchez",slug:"norma-francenia-santos-sanchez",fullName:"Norma Francenia Santos-Sánchez",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/148546/images/4640_n.jpg",biography:null,institutionString:null,institution:null},{id:"272889",title:"Dr.",name:"Narendra",middleName:null,surname:"Maddu",slug:"narendra-maddu",fullName:"Narendra Maddu",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/272889/images/10758_n.jpg",biography:null,institutionString:null,institution:null},{id:"242491",title:"Prof.",name:"Angelica",middleName:null,surname:"Rueda",slug:"angelica-rueda",fullName:"Angelica Rueda",position:"Investigador Cinvestav 3B",profilePictureURL:"https://mts.intechopen.com/storage/users/242491/images/6765_n.jpg",biography:null,institutionString:null,institution:null},{id:"88631",title:"Dr.",name:"Ivan",middleName:null,surname:"Petyaev",slug:"ivan-petyaev",fullName:"Ivan Petyaev",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"Lycotec (United Kingdom)",country:{name:"United Kingdom"}}},{id:"423869",title:"Ms.",name:"Smita",middleName:null,surname:"Rai",slug:"smita-rai",fullName:"Smita Rai",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"Integral University",country:{name:"India"}}},{id:"424024",title:"Prof.",name:"Swati",middleName:null,surname:"Sharma",slug:"swati-sharma",fullName:"Swati Sharma",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"Integral University",country:{name:"India"}}},{id:"439112",title:"MSc.",name:"Touseef",middleName:null,surname:"Fatima",slug:"touseef-fatima",fullName:"Touseef Fatima",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"Integral University",country:{name:"India"}}},{id:"424836",title:"Dr.",name:"Orsolya",middleName:null,surname:"Borsai",slug:"orsolya-borsai",fullName:"Orsolya Borsai",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"University of Agricultural Sciences and Veterinary Medicine of Cluj-Napoca",country:{name:"Romania"}}},{id:"422262",title:"Ph.D.",name:"Paola Andrea",middleName:null,surname:"Palmeros-Suárez",slug:"paola-andrea-palmeros-suarez",fullName:"Paola Andrea Palmeros-Suárez",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"University of Guadalajara",country:{name:"Mexico"}}}]}},subseries:{item:{id:"12",type:"subseries",title:"Human Physiology",keywords:"Anatomy, Cells, Organs, Systems, Homeostasis, Functions",scope:"Human physiology is the scientific exploration of the various functions (physical, biochemical, and mechanical properties) of humans, their organs, and their constituent cells. The endocrine and nervous systems play important roles in maintaining homeostasis in the human body. Integration, which is the biological basis of physiology, is achieved through communication between the many overlapping functions of the human body's systems, which takes place through electrical and chemical means. Much of the basis of our knowledge of human physiology has been provided by animal experiments. Because of the close relationship between structure and function, studies in human physiology and anatomy seek to understand the mechanisms that help the human body function. The series on human physiology deals with the various mechanisms of interaction between the various organs, nerves, and cells in the human body.",coverUrl:"https://cdn.intechopen.com/series_topics/covers/12.jpg",hasOnlineFirst:!0,hasPublishedBooks:!0,annualVolume:11408,editor:{id:"195829",title:"Prof.",name:"Kunihiro",middleName:null,surname:"Sakuma",slug:"kunihiro-sakuma",fullName:"Kunihiro Sakuma",profilePictureURL:"https://mts.intechopen.com/storage/users/195829/images/system/195829.jpg",biography:"Professor Kunihiro Sakuma, Ph.D., currently works in the Institute for Liberal Arts at the Tokyo Institute of Technology. He is a physiologist working in the field of skeletal muscle. He was awarded his sports science diploma in 1995 by the University of Tsukuba and began his scientific work at the Department of Physiology, Aichi Human Service Center, focusing on the molecular mechanism of congenital muscular dystrophy and normal muscle regeneration. His interest later turned to the molecular mechanism and attenuating strategy of sarcopenia (age-related muscle atrophy). His opinion is to attenuate sarcopenia by improving autophagic defects using nutrient- and pharmaceutical-based treatments.",institutionString:null,institution:{name:"Tokyo Institute of Technology",institutionURL:null,country:{name:"Japan"}}},editorTwo:null,editorThree:{id:"331519",title:"Dr.",name:"Kotomi",middleName:null,surname:"Sakai",slug:"kotomi-sakai",fullName:"Kotomi Sakai",profilePictureURL:"https://s3.us-east-1.amazonaws.com/intech-files/0033Y000031QtFXQA0/Profile_Picture_1637053227318",biography:"Senior researcher Kotomi Sakai, Ph.D., MPH, works at the Research Organization of Science and Technology in Ritsumeikan University. She is a researcher in the geriatric rehabilitation and public health field. She received Ph.D. from Nihon University and MPH from St.Luke’s International University. 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