Display Settings:

Format

Send to:

Choose Destination
See comment in PubMed Commons below
J Immunol Methods. 2012 Aug 31;382(1-2):117-28. doi: 10.1016/j.jim.2012.05.011. Epub 2012 May 22.

Optimization and qualification of a multiplex bead array to assess cytokine and chemokine production by vaccine-specific cells.

Author information

  • 1Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Research Center, Seattle, WA 98109, USA.

Abstract

The magnitude and functional phenotype (e.g. proliferation, immune stimulation) of vaccine-induced T-cell responses are likely to be critical in defining responses that can control pathogenic challenge. Current multi-parameter flow cytometric techniques may not be sufficient to measure all of these different functions, since characterizing T-cell responses by flow cytometry is presently limited to concurrent measurement of at most 10 cytokines/chemokines. Here, we describe extensive studies conducted using standardized GCLP procedures to optimize and qualitatively/quantitatively qualify a multiplex bead array (MBA) performed on supernatant collected from stimulated peripheral blood mononuclear cells (PBMC) to assess 12 cytokines and chemokines of interest. Our optimized MBA shows good precision (intra-assay, inter-day, inter-technician; coefficients of variation <30%) and linearity for most of the analytes studied. We also developed positivity criteria that allow us to define a response as positive or negative with a high degree of confidence. In conclusion, we provide a detailed description of the qualification of an MBA, which permits quantitative and qualitative evaluation of vaccine-induced immunogenicity and analysis of immune correlates of protection. This assay provides an excellent complement to the existing repertoire of assays for assessing immunogenicity in HIV vaccine clinical trials.

Copyright © 2012 Elsevier B.V. All rights reserved.

PMID:
22626638
[PubMed - indexed for MEDLINE]
PMCID:
PMC3406745
Free PMC Article

Images from this publication.See all images (7)Free text

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
Figure 7
PubMed Commons home

PubMed Commons

0 comments
How to join PubMed Commons

    Supplemental Content

    Full text links

    Icon for Elsevier Science Icon for PubMed Central
    Loading ...
    Write to the Help Desk