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Updated: Aug 8, 2026
Visualizing Antigen Specific CD4+ T Cells using MHC Class II Tetramers
Published on: March 6, 2009
Definition of an immunologic response using the major histocompatibility complex tetramer and enzyme-linked
Begoña Comin-Anduix1, Antonio Gualberto, John A Glaspy
1Department of Surgery, Division of Surgical, University of California at Los Angeles, Los Angeles, California, USA.
Purpose:
Define an immunologic response using the tetramer and enzyme-linked immunospot (ELISPOT) assays.
Experimental Design:
Ten healthy subjects and 21 patients with melanoma (all HLA-A*0201) donated a total of 121 blood samples to determine the lower limit of detection (LLD), analytic coefficient of variation (aCV), and physiologic CV (pCV) of the tetramer and ELISPOT assays. The mean, SD, and reference change value (RCV) were calculated to define changes beyond the assay imprecision, and its application was tested in the monitoring of T-cell expansion after CTLA4 blockade with ticilimumab (CP-675,206).
Results:
The LLD for the tetramer assay was 0.038% CD8+ cells and seven spots per 10(5) peripheral blood mononuclear cells for the ELISPOT assay. The aCV of the tetramer assay was <10% and was higher for the ELISPOT (24.69-36.32%). There was marked between-subject variability on baseline homeostatic values, which was correlated to prior antigen exposure. An immunologic response was defined as an increase beyond the mean + 3 SD in antigen-specific cells for subjects with baseline levels below the LLD, or beyond the assay RCV for baseline levels above the LLD. In four patients receiving ticilimumab, expansions of antigen-specific T cells beyond the assay variability were noted for EBV and MART1 antigens.
Conclusions:
A combined approach of change from negative (below the LLD) to positive (above the LLD) and a percentage change beyond the assay variability using the RCV score can be computed to define which change in circulating antigen-specific T cells represents a response to immunotherapy.
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