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Enzyme-Linked Immunosorbent Assay

Enzyme-Linked Immunosorbent Assay

In 1971, Peter Perlman and Eva Engvall developed an Enzyme-linked immunosorbent assay (ELISA or EIA). ELISA differs from western blot in that the assays are conducted in microtiter plates or in vivo rather than on an absorbent membrane.
There are many different types of ELISAs, but they all involve an antibody molecule whose constant region binds an enzyme, leaving the variable region free to bind its specific antigen. Enzyme-substrate reaction allows the antigen to be visualized or quantified.

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Related Experiment Video

Updated: Jun 12, 2026

Sigma's Non-specific Protease Activity Assay - Casein as a Substrate

Sigma's Non-specific Protease Activity Assay - Casein as a Substrate

Published on: September 17, 2008

ELISA kit for casein determination: interlaboratory study.

Frantisek Stumr¹, Dana Gabrovská, Jana Rysová

¹SEDIUM RD, Ltd, Zeleznicního pluku 1361, 530 02 Pardubice, Czech Republic.

Journal of AOAC International

|May 20, 2010

Summary

An interlaboratory study validated an ELISA kit for quantifying casein in foods. This enzyme-linked immunosorbent assay method proved specific and reliable for detecting milk protein in various food products.

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Sigma's Non-specific Protease Activity Assay - Casein as a Substrate

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Area of Science:

Food Science
Analytical Chemistry
Immunology

Background:

Accurate quantification of casein is crucial for food labeling and allergen control.
Existing methods may lack specificity or require complex procedures.

Purpose of the Study:

To validate an enzyme-linked immunosorbent assay (ELISA) kit for the quantitative determination of casein in diverse food matrices.
To assess the specificity, limit of quantification (LOQ), and limit of detection (LOD) of the ELISA kit.

Main Methods:

An interlaboratory study involving eight laboratories.
Validation of an ELISA kit utilizing rabbit polyclonal antibodies.
Testing of nine different food samples, including those with and without declared milk ingredients.

Main Results:

The ELISA kit demonstrated high specificity, with no false positives or cross-reactivities observed.
Casein content was below the limit of quantification (LOQ) in most samples without milk proteins.
The kit accurately detected casein in samples containing milk, with levels varying based on declared ingredients. Calculated LOQ: 1.8 mg CAS/kg, LOD: 0.5 mg CAS/kg.

Conclusions:

The validated ELISA method is suitable for the specific and reliable quantification of casein in a wide range of food products.
The method provides accurate results for both milk-free and milk-containing foods, aiding in allergen management and regulatory compliance.