Unit

UNIT 6.19 ELISPOT Assay to Detect Cytokine-Secreting Murine and Human Cells

  1. Dennis M. Klinman1,
  2. Thomas B. Nutman2

Published Online: 1 MAY 2001

DOI: 10.1002/0471142735.im0619s10

Current Protocols in Immunology

Current Protocols in Immunology

How to Cite

Klinman, D. M. and Nutman, T. B. 2001. ELISPOT Assay to Detect Cytokine-Secreting Murine and Human Cells. Current Protocols in Immunology. 10:6.19:6.19.1–6.19.8.

Author Information

  1. 1

    Center for Biologics Evaluation & Research, Food and Drug Administration, Bethesda, Maryland

  2. 2

    National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland

Publication History

  1. Published Online: 1 MAY 2001
  2. Published Print: JUN 1994

This is not the most recent version of the article. View current version (1 NOV 2008)

Abstract

The filter immunoplaque assay, otherwise called the enzyme-linked immunospot assay (ELISPOT), was initially developed to detect and quantitate individual antibody-secreting B cells. Recent modifications have improved the sensitivity of the ELISPOT assay such that cells producing as few as 100 molecules of specific protein per second can be detected. The ELISPOT assay utilizes two high-affinity cytokine-specific antibodies directed against different epitopes on the same cytokine molecule: either two monoclonal antibodies or a combination of one monoclonal antibody and one polyvalent antiserum. ELISPOT generates spots based on a colorimetric reaction that detects the cytokine secreted by a single cell. The spot represents a “footprint” of the original cytokine-producing cell. Spots are permanent and can be quantitated visually, microscopically, or electronically.