Chemical inactivation of recombinant vaccinia viruses and the effects on antigenicity and immunogenicity of recombinant simian immunodeficiency virus envelope glycoproteins.

The efficiency of paraformaldehyde (PFA) and binary ethylenimine (BEI) in inactivating recombinant vaccinia virus (rVV), present in baby hamster kidney cells expressing simian immunodeficiency virus envelope glycoproteins (SIV-Env), was measured in a series of inactivation studies. Both compounds were shown to be effective in reducing rVV titres. The use of standard 3-day titration assays proved to be inadequate to measure PFA inactivation, since upon prolonged incubation, residual rVV infectivity was detected in cultures negative at 3 days. Different procedures using PFA or BEI were selected to assess their influence on the antigenicity and immunogenicity or rVV expressed SIV-Env. Antigenicity, as defined by the ability to react with a panel of monoclonal antibodies recognizing major antigenic sites, and immunogenicity, as defined by the ability to induce SIV envelope specific and virus neutralizing serum antibodies in rats, proved to be preserved after either inactivation procedure. These data show that both protocols using

Simian immunodeficiency virus (SIV)-specific CD8+ cytotoxic T lymphocyte responses of naive and vaccinated cynomolgus macaques infected with (SIV)mac32H(J5): quantitative analysis by in vitro antigenic stimulation.

Detailed analyses of simian immunodeficiency virus (SIV)-specific cytotoxic T lymphocyte (CTL) responses in vaccinated and infected macaques may help to clarify the role of CTL immunity in protection against lentiviruses. Here, the optimal conditions for the measurement of SIV Gag-specific CTL were investigated by bulk and limiting dilution assays of peripheral blood mononuclear cells (PBMC) from naive and vaccinated cynomolgus macaques (Macaca fascicularis) infected with SIVmac32H(J5). In vitro restimulation was generally required for CTL detection. Selective activation of CD8+ and MHC-restricted SIV Gag-specific CTL was induced by stimulation with autologous para-formaldehyde-fixed B-lymphoblastoid cell lines infected with a recombinant vaccinia virus expressing SIV Gag. Applied to limiting dilution assays, antigenic stimulation reproducibly demonstrated SIV Gag-specific CTL precursors (CTLp) in PBMC of all animals studied, including those lacking significant responses in standard bulk CTL assays

Decline of simian immunodeficiency virus (SIV)-specific cytotoxic T lymphocytes in the peripheral blood of long-term nonprogressing macaques infected with SIVmac32H-J5.

The evolution of simian immunodeficiency virus (SIV)–specific cytotoxic T lymphocyte precursors (CTLps) and their relationship with virus replication were studied in SIV‐infected macaques. After primary viremia, 3 of 8 macaques l

CD8+ cytotoxic T lymphocytes of a cynomolgus macaque infected with simian immunodeficiency virus (SIV)mac32H-J5 recognize a nine amino acid epitope in SIV Gag p26.

A detailed analysis of simian immunodeficiency virus (SIV)-specific cytotoxic T lymphocyte (CTL) responses and the identification of the proteins and epitopes they target may improve the design of immunotherapeutic interventions and provide insights into AIDS pathogenesis. Here, we identified a new CTL epitope in the SIV Gag protein, recognized by CD8+ and MHC class I-restricted CTL clones from a long-term asymptomatic cynomolgus macaque (Macaca fascicularis) infected with SIVmac32H-J5. Using overlapping synthetic peptides, the optimal minimal epitope was characterized as a nine amino acid peptide representing amino acids 242-250 of p26 (SVDEQIQWM). CTL recognition was shown to be abolished by amino acid substitutions observed within homologous human immunodeficiency virus (HIV)-1 and HIV-2 sequences

Comparison of protection from homologous cell-free vs cell-associated SIV challenge afforded by inactivated whole SIV vaccines.

This study attempted to determine if SIV vaccines could protect against challenge with peripheral blood mononuclear cells (PBMCs) from an SIV infected rhesus monkey. Mature Macaca mulatta were vaccinated four times with formalin inactivated SIVmac32H administered in MDP adjuvant (n = 8) or SIVmac32H ISCOM vaccine (n = 8). Controls included animals vaccinated with measles virus in MDP adjuvant (n = 4) or ISCOM (n = 4) preparations. Of each group, half were challenged intravenously (IV) with ten MID50 of the cell-free SIVmac32H (11-88) SIV stock and half were challenged with ten MID50 of PBMCs from the SIVmac32H infected macaque 1XC. All SIV vaccinated animals challenged with the 11-88 cell free stock of SIVmac32H were protected, whereas only half of the SIV vaccinated monkeys receiving the same infectious dose of the 1XC cell stock were protected