ELISA Guidebook.pdf

The ELISA Guidebook

Second Edition

Series Editor

John M. Walker

School of Life Sciences

University of Hertfordshire

Hatfield, Hertfordshire, AL10 9AB, UK

For other titles published in this series, go to

www.springer.com/series/7651

M ETHODS IN M OLECULAR B IOLOGY ™

The ELISA Guidebook

Second Edition

John R. Crowther

International Atomic Energy Agency, Animal Production & Health Section, Vienna, Austria

John R. Crowther

International Atomic Energy Agency

Animal Production & Health Section

Wagramer Str. 5

1400 Vienna

Austria

ISBN: 978-1-60327-253-7

e-ISBN: 978-1-60327-254-4

ISSN: 1064-3745

e-ISSN: 1940-6029

DOI: 10.1007/978-1-60327-254-4

Library of Congress Control Number: 2008940983

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Preface

There have been very few developments that markedly affect the need to greatly revise

the text from the last version of this book. This is testament to the fact that heteroge-

neous enzyme-linked immunosorbent assays (ELISA) provide ideal systems for dealing

with a wide range of studies in many biological areas. The main reason for this success is

test flexibility, whereby reactants can be used in different combinations, either attached

passively to a solid phase support or in the liquid phase. The exploitation of the ELISA

has been increased through continued development of specifically produced reagents, for

example, monoclonal and polyclonal antibodies and peptide antigens coupled with the

improvement and expansion of commercial products such as enzyme-linked conjugates,

substrates and chromogens, plastics technology and design of microwell plates, instru-

mentation advances and robotics. However, the principles of the ELISA remain the same.

There has been some rearrangement of chapters plus addition of three new ones dealing

with charting methods for assessing the indirect ELISA, ruggedness and robustness of

tests-aspects of kit use and validation, and internal quality control and external quality

management of data, respectively. These reflect the need to control what you are doing

with ELISA and to exploit the method to its full extent. I do not apologize for dealing

with the same areas in different ways a number of times, as it is imperative that principles

are understood to allow planning, operation, and control of ELISA.

A brief scan of the literature involving ELISA can be used to illustrate the contin-

ued success of ELISA. The number of publications with ELISA mentioned in all science

areas from 1976 to 2004 is shown in Table 1. A fairly constant increase in the number of

research works using ELISA methods is indicated. A breakdown of publications accord-

ing to the areas of science in 5 yearly periods from 1980 given in Table 2 illustrates the

versatility in the use of ELISA, as well as highlights the major areas of use in medicine and

dentistry; immunology and microbiology, molecular biology, and genetics and biotech-

nology. It is interesting to note that the earliest exploitation of ELISA was in immunol-

ogy and microbiology and molecular Biology and biotechnology, probably reflecting the

greatest research areas. Medicine and dentistry (associated by the search engine) shows

the greatest rate of increase in use (probably in the medical sphere only) from the 1990s.

The search results indicate the continued expansion of ELISA in science, and there is

no reason to believe that this will change even in the face of modern technologies exploit-

ing molecular methods. The analytical and systematic characteristics of ELISA are ideally

suited to diagnosis at the screening level, for surveillance where larger scale sample handling

is required, and for research. Many of the accepted standard assays in many scientific

fields are ELISA-based and have replaced other “gold standard” assays. In conjunction

with the rapidly evolving use of molecular methods centering on the polymerase chain

reaction (PCR) technologies, there is a need to use serological confirmatory methods in

a dual approach to directly identify and characterize disease agents and to assess disease

prevalence through the measurement of specific antibodies or other chemical factors as

a result of infection. The use of ELISA methods in testing the environment and animal

or plant products as safe for human and animal consumption is also a rapidly evolving

area for ELISA.

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