Abstract
The enzyme-linked immunosorbent assay (ELISA) is a simple and rapid technique for detecting and quantitating antibodies or antigens attached to a solid surface. Being one of the most sensitive immunoassays, ELISA offers commercial value in laboratory research, diagnostic of disease biomarkers, and quality control in various industries. This technique utilizes an enzyme-linked antibody binding to a surface-attached antigen. Subsequently, a substrate is added to produce either a color change or light signal correlating to the amount of the antigen present in the original sample. This chapter provides the procedures required for carrying out indirect ELISA, one of the many forms of ELISA, to detect polystyrene-immobilized antigen. Methodological approaches to optimize this assay technique are also described, a prerequisite for automation and multiplexing.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Engvall E, Perlmann P (1971) Enzyme-linked immunosorbent assay (ELISA). Quantitative assay of immunoglobulin G. Immunochemistry 8:871–874
Stevens PW, Hansberry MR, Kelso DM (1995) Assessment of adsorption and adhesion of proteins to polystyrene microwells by sequential enzyme-linked immunosorbent assay analysis. Anal Biochem 225:197–205
Desmet C, Blum LJ, Marquette CA (2013) Multiplex microarray ELISA versus classical ELISA, a comparison study of pollutant sensing for environmental analysis. Environ Sci Process Impacts 10:1876–1882
Fan A, Cao Z, Li H et al (2009) Chemiluminescence platforms in immunoassay and DNA analyses. Anal Sci 25:5875–5897
Gan SD, Patel KR (2013) Enzyme immunoassay and enzyme-linked immunosorbent assay. J Invest Dermatol 133:e12
Hornbeck P (2001) Enzyme-linked immunosorbent assays. Curr Protoc Immunol Chapter 2: Unit 2.1. doi: 10.1002/0471142735.im0201s01
Natarajan S, Remick DG (2008) The ELISA standard save: calculation of sample concentrations in assays with a failed standard curve. J Immunol Methods 336:242–245
Underwood PA, Steele JG (1991) Practical limitations of estimation of protein adsorption to polymer surfaces. J Immunol Methods 142:83–94
Hnasko R, Lin A, McGarvey JA et al (2011) A rapid method to improve protein detection by indirect ELISA. Biochem Biophys Res Commun 410:726–731
Vos Q, Klasen EA, Haaijman JJ (1987) The effect of divalent and univalent binding on antibody titration curves in solid-phase ELISA. J Immunol Methods 103:47–54
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2015 Springer Science+Business Media New York
About this protocol
Cite this protocol
Lin, A.V. (2015). Indirect ELISA. In: Hnasko, R. (eds) ELISA. Methods in Molecular Biology, vol 1318. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2742-5_5
Download citation
DOI: https://doi.org/10.1007/978-1-4939-2742-5_5
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-2741-8
Online ISBN: 978-1-4939-2742-5
eBook Packages: Springer Protocols