The Amplified Enzyme-Linked Immunosorbent Assay (a-ELISA)

  • J. E. Butler
  • J. H. Peterman
  • T. E. Koertge


The current interest in immune regulation and in characterizing the immune response against pathogens and substances of interest to human and animal medicine, has prompted attempts to develop techniques capable of accurately and reliably measuring specific antibodies in serum and body fluids. Especially important is the ability to measure the response according to the isotype of the antibody. The enzyme-linked immunosorbent assay (ELISA) provides a sensitive and non-hazardous immunochemical basis for potentially making such determinations (Avrameas and Gilbert, 1972; Engvall et al. 1971). The amplified ELISA (a-ELISA) is a modification of the original ELISA which combines the principles of indirect detection with the use of a non-covalent enzyme-antibody detection system (Butler et al. 1978b). The latter avoids alteration of both enzyme and antibody, the complex in fact dissociating during the substrate reaction. The result is a system with improved sensitivity and broad flexibility. The a-ELISA serves as a useful means of measuring the isotypic distribution of specific immune responses in many species.


Titration Curve Linear Region Antibody Affinity ELISA Titration Dilution Sequence 
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  1. Axen, R. and P. Vretblad. Chemical fixation of proteins to water-insoluble carriers, in: Protides of the Biological Fluids. Vol. 18. H. Peeters, ed., Pergamon Press, Oxford (1971).Google Scholar
  2. Avrameas, S. A. and B. Guilbert. Enzyme-immunoassay for the measurement of antigens using peroxidase conjugates. Biochimie 54:837 (1972).PubMedCrossRefGoogle Scholar
  3. Bruni, C, A. Germani, G. Koch and R. Strom. Derivation of antibody distribution from experimental binding data. J. Theor. Biol. 61:143 (1976).PubMedCrossRefGoogle Scholar
  4. Butler, J. E. The amplified ELISA: Principles and applications for the comparative quantitation of class and subclass antibodies and the distribution of antibodies and antigens in biochemical separates. Methods in Enzymology, Vol. 73, Immunochemical Methods. H. J. Vunakis and J. J. Lagone, ed. (1981).Google Scholar
  5. Butler, J. E., T. L. Feldbush, P. L. McGivern and N. Stewart. The enzyme-linked immunosorbent assay (ELISA): A measure of antibody concentration or affinity? Immunochemistry 15:131 (1978a).PubMedCrossRefGoogle Scholar
  6. Butler, J. E., P. L. McGivern and P. Swanson. Amplification of the enzyme-linked immunosorbent assay (ELISA) in the detection of class-specific antibodies. J. Immunol. Methods 20:365 (1978b).PubMedCrossRefGoogle Scholar
  7. Butler, J. E., and P. L. McGivern. The measurement of IgE and other classes of antibodies in the sera and bronchial lavage fluids of patients to Alternaría, house dust and ragweed. (1985, Submitted).Google Scholar
  8. Butler, J. E., P. L. McGivern, L. A. Cantarero, and L. Peterson. Application of the amplified enzyem-linked immunosorbent assay: Comparative quantitation of bovine serum IgGl, IgG2, IgA and IgM antibodies. Am. J. Vet. Res. 41:1479 (1980a).PubMedGoogle Scholar
  9. Butler, J. E., L. Peterson and P. L. McGivern. A reliable method for the preparation of bovine secretory immunoglobulin A (SIgA) which circumvents problems posed by IgGl dimers in colostrum. Mol. Immunol. 17:757 (1980b).PubMedCrossRefGoogle Scholar
  10. Cantarero, L., J. E. Butler and J. W. Osbourne. The binding characteristics of six proteins to polystyrene and its implications to solid-phase immunoassays using polystyrene tubes. Analy. Biochem. 105:375 (1980).Google Scholar
  11. Cuatrecasas, P., and C. B. Anfinsen. Affinity chromatography.Annual Rev. Biochem. 40:259 (1971).CrossRefGoogle Scholar
  12. Dierks, S. E. Absolute quantitation of IgG and IgA anti-protein and anti-hapten antibodies using a microELISA system. M.S. Thesis, University of Iowa (1985).Google Scholar
  13. Engvall, E., K. Jonsson and P. Perlmann. Enzyme-linked immunosorbent assay. II. Quantitative assay of protein antigen, immunoglobulin G, by means of enzyme labelled antigen and antibody coated tubes. Biochim. Biophys. Acta. 251:472 (1971).Google Scholar
  14. Engvall, E., and P. Perlmann. Enzyme-linked immunosorbent assay, ELISA. III. Quantitation of specific antibodies by enzyme-labelled anti-immunoglobulin in antigen-coated tubes. J. Immunol. 109:129 (1972).PubMedGoogle Scholar
  15. Koertge, T. E. A study of the quantitative capability of the enzyme-linked immunosorbent assay (ELISA) with regard to the use of specific antibodies to study the transport of monomeric and polymeric antibodies. Ph.D. Thesis, University of Iowa (1984).Google Scholar
  16. Koertge, T. E., and J. E. Butler. The relationship between the binding of primary antibody to solid-phase antigen in microtiter plates and its detection by the ELISA. J. Immunol. Methods (1985, submitted).Google Scholar
  17. Koertge, T. E., J. E. Butler and S. E. Dierks. Evaluation of the soluble immune complex (EIC) of the amplified enzyme-linked immunosorbent assay (a-ELISA) and the use of this assay for quantitation by reaction stoichiometry (1985, submitted).Google Scholar
  18. Lehtonen, O.-P., and E. Eerola. The effect of different antibody affinities on ELISA absorbance and titer. J. Immunol. Methods 54:233 (1982).PubMedCrossRefGoogle Scholar
  19. Lew, A. M. The effect of epitope density and antibody affinity on the ELISA as analyzed by monoclonal antibodies. J. Immunol. Methods 72:171 (1984).PubMedCrossRefGoogle Scholar
  20. Nimmo, G. R., A. M. Lew, C. M. Stanley and M. W. Steward. Influence of antibody affinity on the performance of different antibody assays. J. Immunol. 72:177 (1984).Google Scholar
  21. Peterfy, F., P. Kunsela and O. Makela. Affinity requirements for antibody assays mapped by monoclonal antibodies. J. Immunol. 130:1809 (1983).PubMedGoogle Scholar
  22. Peterman, J. H. The influence of antibody affinity, valency and heterogeneity on binding behavior of antifluorescein antibodies in solid-phase micro-ELISA systems. Ph.D. Thesis, University of Iowa (1985).Google Scholar
  23. Pringnitz, D. J., and J. E. Butler. Bovine-associated muco-protein (BAMP). III. De novo synthesis by non-mammary tissues. J. Dairy Sci. (1985, in press).Google Scholar
  24. Pringnitz, D. J., J. E. Butler and A. J. Guidry. In vivo proleolytic activity of the mammary gland. Contribution to the origin of secretory component, β2-microglobulin and bovine-associated mucoprotein (BAMP; in cows milk. Vet. Immunol. Immunopath. (1985a, in press).Google Scholar
  25. Pringnitz, D. J., J. E. Butler and A. J. Guidry. Quantitation of bovine β2-microglobulin: Occurrence in body fluids, on milk fat globules and origin in milk. Mol. Immunol. (1985b, in press).Google Scholar
  26. Sips, R. On the structure of the catalyst surface. J. Chem. Phys. 16:490 (1948).CrossRefGoogle Scholar
  27. Sloan, G. J. Enzyme-linked immunosorbent assay (ELISA): Application to the quantitation by subclass of bovine antibodies against Staphylococcus aureus. M.S. Thesis, University of Iowa (1975).Google Scholar
  28. Yorde, D. E., E. A. Sasse, T. Y. Wang, R. O. Hussa and J. C. Garancis. Competitive enzyme-linked immunoassay with use of a soluble enzyme/antibody immune complex for labeling. I. Measurement of human choriogonadotropin. Clin. Chem. 22:1372 (1976).PubMedGoogle Scholar
  29. Zeiss, C. R., J. J. Pruzansky, R. Patterson and M. Roberts. A solid phase radioimmunoassay for the quantitation of human reaginic antibody against ragweed antigen E. J. Immunol. 110:414 (1973).PubMedGoogle Scholar

Copyright information

© Plenum Press, New York 1985

Authors and Affiliations

  • J. E. Butler
    • 1
  • J. H. Peterman
    • 1
  • T. E. Koertge
    • 2
  1. 1.Department of MicrobiologyThe University of Iowa Medical SchoolIowa CityUSA
  2. 2.Department of Periodontics, Medical College of VirginiaVirginia Commonwealth UniversityRichmondUSA

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