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57 Rapid and Comprehensive Characterization of HIV-1-Specific CTL Responses Using a Peptide-Matrix-System Spanning All Expressed HIV-1 Proteins
M. M. Addo1, X. G. Yu*1, D. Cohen2, R. Eldridge1, M. Philips1, C. Brander1, S. Boswell2, P. Goulder1, E. Rosenberg1, M. Altfeld1, and B. D. Walker1
1AIDS Res. Ctr., MGH, Boston, MA, USA and 2Fenway CHC, Boston, MA, USA
Background: Cellular immune responses play a critical role in the control of HIV-1. However, no comprehensive analysis of HIV-1-specific T-lymphocyte responses directed against the whole expressed genome has been performed. This study evaluates a rapid peptide-matrix-based screening approach requiring limited input cell numbers and also represents the first report of comprehensive screening of CD8 T-cell responses on an epitope level in HIV-1-infected individuals.
Methods: PBMC from 30 HIV-1-infected individuals were screened for HIV-specific T-cell responses by Elispot using a peptide-matrix with 105 peptide pools including 505 overlapping peptides spanning all expressed HIV-1 proteins. In parallel, the 505 overlapping peptides were individually tested. An evaluation of specificity and sensitivity of the method was performed and total breadth and magnitude of T-cell responses in each individual were assessed.
Results: Compared with screening with individual overlapping peptides, the peptide matrix approach showed a sensitivity of 91% (98% for responses >300 SFC/106 PBMC) and a specificity of 98%. Magnitudes of responses to the peptide matrix correlated well to screening with individual peptides. The matrix-based approach reduced the amount of PBMC needed for a complete screening against the entire expressed HIV-1 genome by up to 80%. Gag was most frequently targeted followed by Pol and Nef. The number of total recognized HIV-1 peptides ranged from 3 to 56 (median 17), the total magnitude of peptide responses ranged from 290 to >15,000 SFC/106 PBMC.
Conclusions: Using a peptide matrix system allows for the rapid screening of lymphocyte responses directed against the whole expressed HIV-1 genome with high sensitivity and specificity. The amount of required cells, cost and time are significantly reduced compared with screening with individual peptides, which makes this method a valuable tool for the assessment of HIV-1-specific cellular immune responses in large scale immunotherapy and vaccine trials. This study also demonstrates that virus-specific immune responses are directed against a large number of different epitopes.
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