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Related Concept Videos

Enzyme-Linked Immunosorbent Assay01:33

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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High Throughput Sequential ELISA for Validation of Biomarkers of Acute Graft-Versus-Host Disease
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Multisubstrate-compatible ELISA procedures for rapid and high-sensitivity immunoassays.

Chandra Kumar Dixit1, Sandeep Kumar Vashist, Brian D MacCraith

  • 1Centre for Bioanalytical Sciences, National Centre for Sensor Research, Dublin City University, Dublin, Ireland.

Nature Protocols
|March 18, 2011
PubMed
Summary

This study presents a novel method for developing highly sensitive enzyme-linked immunosorbent assays (ELISAs) through covalent antibody immobilization. This optimized protocol significantly enhances assay sensitivity and speed for various applications.

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An ELISA Based Binding and Competition Method to Rapidly Determine Ligand-receptor Interactions
08:40

An ELISA Based Binding and Competition Method to Rapidly Determine Ligand-receptor Interactions

Published on: March 14, 2016

Area of Science:

  • Biochemistry and Molecular Biology
  • Analytical Chemistry
  • Materials Science

Background:

  • Enzyme-linked immunosorbent assays (ELISAs) are crucial for detecting and quantifying analytes.
  • Existing ELISA methods often face limitations in sensitivity and assay duration.
  • Optimizing antibody immobilization is key to improving ELISA performance.

Purpose of the Study:

  • To develop an improved and optimized protocol for rapid and high-sensitivity ELISAs.
  • To achieve covalent antibody immobilization on chemically modified polymeric surfaces.
  • To enhance the sensitivity and speed of ELISA procedures for clinical and research applications.

Main Methods:

  • Surface activation of polymeric materials using potassium hydroxide (KOH) and oxygen (O2) plasma.
  • Amine functionalization of the activated surface using 3-aminopropyltriethoxysilane.
  • Covalent immobilization of antibodies onto the aminated surface, followed by ELISA development.

Main Results:

  • The developed ELISA protocol demonstrated a 16-fold increase in sensitivity compared to established methods.
  • The antibody immobilization procedure requires approximately 3 hours, enabling rapid ELISAs.
  • The method is applicable to a wide range of solid support matrices, including chemically inert materials.

Conclusions:

  • This protocol offers a generalizable quantitative analytical approach for high-sensitivity and rapid assays.
  • The method is suitable for clinical diagnostics and accelerates the screening of phage-displayed libraries.
  • The covalent antibody immobilization technique enhances ELISA performance across various biosensor formats.