TAg

Antigen-specific T lymphocytes must be quantified in order to gauge the quality of an immune response. Typically this is accomplished using cytotoxicity assays or limiting dilution analysis (LDA), but these techniques are lengthy and provide indirect quantitation. Also, LDA cannot count nonproliferative cells. In 1996, Stanford University's Mark Davis developed an alternative strategy that overcomes these problems.1 Davis generated phycoerythrin-conjugated tetramers of human lymphocyte antigen (

Apr 30, 2001
Jeffrey Perkel
Antigen-specific T lymphocytes must be quantified in order to gauge the quality of an immune response. Typically this is accomplished using cytotoxicity assays or limiting dilution analysis (LDA), but these techniques are lengthy and provide indirect quantitation. Also, LDA cannot count nonproliferative cells. In 1996, Stanford University's Mark Davis developed an alternative strategy that overcomes these problems.1 Davis generated phycoerythrin-conjugated tetramers of human lymphocyte antigen (HLA)-A2 coupled to specific peptides, and used these tetramers to accurately count CD8+ T cells directed against three specific viral peptides by flow cytometry. Stanford has now licensed the technology to Beckman-Coulter Inc. of Fullerton, Calif., which markets it as i·TAg MHC tetramers.

Beckman-Coulter's new San Diego-based Immunomics Operations currently offers a custom tetramer service as well as standard i·TAgs directed against HIV-1 Gag or Pol, and the melanoma MART-1 peptide. For custom tetramers, researchers provide an amino acid sequence and Immunomics synthesizes an i·TAg containing this peptide. According to Immunomics' marketing director Susan Gammon, i·TAg technology is unique in that it is "really the first technology available that allows you to measure specific populations of T cells." Gammon predicts that i·TAgs will prove most useful in viral, transplantation, and therapeutic cancer vaccine research. For example, i·TAgs could be used as an early predictive measure of transplant rejection, or to determine the efficacy of a treatment designed to boost the immune system's response to cancerous cells.

Markus Maeurer, professor of Medical Microbiology at the University of Mainz, Germany, uses i·TAgs extensively in his research. Initially, Maeurer quantified T-cell responses using indirect methods such as T-cell cloning or flow cytometry of cell populations bound to T-cell receptor-specific antibodies. However, with i·TAgs, he can "look in clinical samples for the number of antigen-specific T cells without the need for ex vivo expansion of T cells," which tends to bias the results. Maeurer notes that one of the primary advantages of i·TAgs is that they allow scientists to "combine the enumeration of antigen-specific T cells with marker analysis or with the detection of intracellular cytokines elicited upon exposure of T cells to their nominal antigen."

Immunomics is currently developing related i·TAg products that will be used to detect antigen-specific helper T cells (CD4+), which recognize antigen presented in MHC class II molecules. According to Gammon, the company is also working to develop new peptide conjugates. The existing i·TAgs detect reactive T cells in those individuals able to recognize antigen in the context of the MHC class I HLA-A2 molecule, which is approximately 40-50 percent of the population.

-Jeffrey M. Perkel (jperkel@the-scientist.com)
1. J.D. Altman, et al., "Phenotypic analysis of antigen-specific T lymphocytes," Science, 274:94-6, 1996.

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