title: Comparison of the ELISPOT and cytokine flow cytometry assays for the enumeration of antigen-specific T cells.
abstract: The enumeration of antigen-specific T cell responses has been greatly facilitated in recent years by the development of methods based on the detection of cytokines. In particular, the enzyme-linked immunospot (ELISPOT) and cytokine flow cytometry (CFC) assays have become popular. Since both assays are likely to continue to be in widespread use, it is important to evaluate whether their results are comparable. In the current study, we compared the results obtained in the ELISPOT and CFC assays using peptide pools corresponding to CMV and HIV-1 proteins in chronically HIV-1-infected individuals. Analysis of T cell responses to peptide pools indicated that the CMV pp65 and HIV-1 Gag CFC and ELISPOT-derived results were statistically correlated. However, the results obtained with each assay differed in important ways: the magnitude of the response was consistently higher in the CFC assay while the CFC assay was less likely than the ELISPOT assay to detect low-level responses. Furthermore, there was a lack of numeric agreement between ELISPOT and CFC results. For studies that require the detection of low-level responses, or definition of responses as positive or negative, the ELISPOT assay may be preferable. In contrast, the CFC has a greater dynamic range and allows for phenotypic discrimination of responding cells, making it the assay of choice for most other applications.
authors.affiliation: Gladstone Institute of Virology and Immunology, University of California-San Francisco, P.O. Box 419100, San Francisco, CA 94141-9100, USA. akarlsson@gladstone.ucsf.edu
authors.forename: Annika C
authors.initials: AC
authors.lastname: Karlsson
authors.forename: Jeffrey N
authors.initials: JN
authors.lastname: Martin
authors.forename: Sophie R
authors.initials: SR
authors.lastname: Younger
authors.forename: Barry M
authors.initials: BM
authors.lastname: Bredt
authors.forename: Lorrie
authors.initials: L
authors.lastname: Epling
authors.forename: Rollie
authors.initials: R
authors.lastname: Ronquillo
authors.forename: Arjun
authors.initials: A
authors.lastname: Varma
authors.forename: Steven G
authors.initials: SG
authors.lastname: Deeks
authors.forename: Joseph M
authors.initials: JM
authors.lastname: McCune
authors.forename: Douglas F
authors.initials: DF
authors.lastname: Nixon
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Sinclair
mesh_terms: D018791:CD4 Lymphocyte Count; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology; D003587:Cytomegalovirus / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005434:Flow Cytometry / Q000379:methods*; D015497:HIV-1 / Q000276:immunology; D006801:Humans; D007371:Interferon-gamma / Q000096:biosynthesis*; D013601:T-Lymphocytes / Q000276:immunology*; D014616:Vaccinia virus / Q000276:immunology
chemical_list: D007371:Interferon-gamma