Methods and devices for preventing transmission of sexually transmitted diseases
a technology of sexually transmitted diseases and prophylaxis, applied in the field of hiv prophylaxis, can solve the problems of inconvenient methods, inability to meet the needs of patients, and inability to follow the method, so as to reduce the risk of transmission and inhibit the effect of transmission
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example i
Preparation of Reagents
A. Specimens
[0037] Seminal fluids are collected from normal males. Samples are stored at 4° C. for 1 to 2 days or alternatively stored at −20° C. for up to three months before use. University of Texas Institutional Review Board Approval Guidelines, including informed consent were followed for all samples collected.
B. Viruses
[0038] Stocks of 213, AC0-1 and 9H strains of HIV were propagated in human H9 lymphocytes using the standard procedure described in Baron et al., Arch. Intern. Med. 159(3):303-310 (1999) and Williams et al., Virology 184(2):723-728 (1991), each of which is incorporated herein by reference. Aliquots were stored frozen at −70° C. The Indiana strain of vesicular stomatitis virus (VSV) was propagated in murine L cells using the method in Coppenhaver et al., New Eng. J. Med. 330(18):1314-1315 (1994), which is incorporated herein by reference. VSV stocks were stored frozen at −70° C.
C. Cells
[0039] Human peripheral blood mononuclear leuko...
example ii
Virus Multiplication
[0040] The effect of various treatments on virus multiplication was determined in virus-infected human CEM lymphocytes, THP-1 macrophages peripheral blood mononuclear leukocytes, or L929 cells. The multiplication of HIV in PBLs was determined as the yield of infectious HIV from 2×106 Ficoll-Hypaque purified normal peripheral blood mononuclear cells that had been cultured with phytopemagglutinin (4 ug / ml) for 2 days before treatment with 20 u / ml of interleukin 2 (40 units / ml) for an additional 2 days and then infected with 105, 50% tissue culture infectious doses of HIV, strain 213. The infected cells were then incubated with 20 u / ml interleukin 2 for five days, washed four times, and then incubated with or without samples, the various mixtures of samples, or culture medium for 15 o 60 minutes. These leukocytes were subsequently washed and cultured for HIV production in RPMI 1640 tissue culture medium, plus 15% fetal bovine serum and 20 u / ml of interleukin 2 for ...
example iii
Assays of HIV and VSV Production
[0041] The production of HIV in the experiments was determined by the standard tissue culture infectious dose 50% assay using MT-2 human lymphocytes, as described in McKeating et al., supra. Specifically, the culture fluids harvested from the HIV-infected human cells were serially diluted in 0.5 log10 increments using RPMI culture medium containing 10% fetal bovine serum. 50 ul of each dilution was added to quadruplicate microtiter wells, each containing 110 ul of 2×104 MT-2 cells in culture medium. After the serial dilutions, 120 ul of nutrient medium was added to each well. These microtiter plates were then incubated at 37° C. for three days in a CO2 incubator, and then re-fed culture medium containing 10% fetal bovine serum. The wells were read for HIV multinucleated giant cell cytopathic effect on day 5 or 6. The 50% tissue culture infections dose was determined using the Reed-Muench method, as described in Reed et al., Am. J. Hyg. 27:493-497 (19...
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