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Direct Detection and Magnetic Isolation of Chlamydia trachomatis Major Outer Membrane Protein-Specific CD8⁺ CTLs with HLA Class I Tetramers¹

Seon-Kyeong Kim^2,3,*, Lesley Devine^2,, Mark Angevine, Robert DeMars^* and Paula B. Kavathas^4,

^* Laboratory of Genetics and Department of Medicine, University of Wisconsin, Madison, WI 53706; and Department of Laboratory Medicine, Yale University School of Medicine, New Haven, CT 06520-8035

We recently identified HLA class I-presented epitopes in the major outer membrane protein (MOMP) of Chlamydia trachomatis that elicit CTL responses in human genital tract infections. T cells possessing cytolytic activities specific for these epitopes could be detected following in vitro stimulation of peripheral blood CD8⁺ T cells with peptides. In the present study we used HLA-A2 tetramers for detailed characterization of MOMP-specific CTL responses. Ex vivo tetramer analysis detected MOMP-specific T cells in the peripheral blood of infected individuals at significant frequencies (0.01–0.20% of CD8⁺ T cells). After in vitro stimulation with peptides, the frequencies of MOMP peptide-specific T cells increased up to 2.34% of CD8⁺ T cells in bulk cultures. In contrast, HLA-A2/MOMP tetramer-binding T cells were virtually undetectable in the peripheral blood from uninfected individuals, either ex vivo or after 3 wk of in vitro peptide stimulation of their T cells. Magnetically sorted, tetramer-bound T cells specifically lysed peptide-pulsed targets as well as C. trachomatis-infected epithelial cells with nearly 50-fold greater per cell efficiency than that of unsorted populations. This study provides conclusive evidence of in vivo induction of HLA class I-restricted CD8⁺ CTL responses to C. trachomatis MOMP. Direct detection of these cells with tetramers will allow their further characterization without prior manipulation and facilitate monitoring of CTL responses during infections and in immunization trials with MOMP-based vaccines.

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