[Evaluation the immunogenesity structure of HIV recombinant protein gp41-p24]

HIV recombinant proteins can be used as immunogen and inducer of immune system. The gag and env proteins are among the most important ones that are located on internal and surface sections, respectively. In this study, we considered the immunogenisity structure of HIV recombinant protein gp41-p24, which has not yet been studied in detail before. In the present study, the HIV recombinant protein gp41-p24 was prepared by cloning methods and kept as unfolded. Using the dilution procedure, unfolded protein was changed to refolded state. The molecular weight and concentration of protein [refolded and unfolded] was measured by electrophoresis and spectrophotometer methods. The measurement of refolded protein can be estimated by native gel and circular dichroism method [CD] based on secondary structure of the protein. Immunological activity and immunogenic structure of these two proteins, based on protein type and optical density was recorded by ELISA and western blot methods. Our results showed that molecular weight of each protein was 32 KD and also they were pure. The refolded protein was observed by native gel method. In the above protein compared with the unfolded one, increased content of helix and beta strand structures and decreased random form was shown. Immune reaction with the antibodies in the serum of HIV positive control patients was observed in the standard and refolded proteins. There was no significant difference based on the protein type. Our research indicated that the HIV recombinant protein gp41-p24, after refolding, has immunogenic activity and we suggest its application as an immunogen in immunization and stimulation of immune system

In vitro susceptibility of Thai seronegative donor CD8+ T lymphocytes to human immunodeficiency virus-1 (HIV-1) infection.

To determine whether CD8+ T lymphocytes from Thai donor cells are susceptible to HIV-1 infection, undepleted peripheral blood mononuclear cells (PBMC) and CD8-enriched PBMC were infected with HIV-1 Thai subtype B and CRF01_AE (E) primary isolates. Virus kinetics in HIV-1 infection of CD4+ and CD8+ T lymphocytes peaked at day 7 or 10 post infection (pi); the TCID50 used for cell infection was proportional to the level of p24 production in the cultures. We also found that the level of p24 antigen in the supernatants of infected undepleted PBMC was significantly higher than that of infected CD8-enriched PBMC. Interestingly, both single positive T lymphocytes (CD4+ and CD8+ T lymphocytes) as well as double positive CD4+/CD8+ T lymphocytes were infected with HIV-1. The double positive T lymphocytes in PBMC were found only in the presence of both CD4+ and CD8+ T lymphocytes. The majority of p24+/CD4-/CD8- T lymphocytes were HIV-1 infected CD4 down-modulated PBMC. This report provides direct evidence that single positive CD8+ T lymphocytes and double positive CD4+/ CD8+ T lymphocytes from Thai donors can be infected with HIV-1 subtypes B and E in vitro.

p24 antigen screening to reduce the risk of HIV transmission by seronegative bone allograft donors.

BACKGROUND: During the last decade, more than 2000 bone allografts harvested from 888 donors and processed by the Queensland Bone Bank have been transplanted in over 1500 patients in Australia and New Zealand. A strict protocol to eliminate HIV transmission by fresh frozen allografts is followed; and not a single case of HIV transmission has been reported. METHODS: All donors were screened and strict donor exclusion criteria were used. All donor blood samples were subjected to double testing including antibody to HIV-1, HIV-2 and HTLV-1 and p24 antigen. The allografts negative for these tests were subjected to processing, including removal of extraneous tissue, pulsatile lavage to remove marrow elements, and immersion in 97% alcohol for 20 minutes. Allografts were subjected to 25 cGy irradiation before transplantation. RESULTS: Allografts were retrieved from a total of 950 donors and 51 were discarded after screening for contamination by organisms other than HIV-1. Eleven donors negative for HIV-1 antibodies tested positive for p24 antigen and were discarded. Allografts from donors testing negative for both the tests (n = 888) were irradiated and used for transplantation. CONCLUSIONS: Routine p24 antigen testing and irradiation of allograft should be mandatory for bone banks, especially those freezing fresh allografts. p24 antigen testing is inexpensive, rapid and easy. Certain guidelines must be followed to avoid misleading results of p24 testing.