Psoralen/UV Inactivation of HIV-1-Infected Cells for Use in Cytologic and Immunologic Procedures

Abstract

A rapid procedure for the inactivation of HIV-1-infected cells using psoralen and ultraviolet (UV) light is described. Exposure of HIV-1-infected cells to 5 micrograms/ml psoralen followed by UV irradiation (320-380 nm) for 5 minutes yields cells that are noninfectious as assessed by extended infectivity assays. The psoralen/UV inactivation procedure described is effective with cells chronically or acutely infected with HIV-1 and is unaffected by cell densities up to 12 x 10(6)/ml. At 5 micrograms/ml psoralen does little damage to cellular permeability as shown by the ability of treated cells to exclude trypan blue and propidium iodide. Psoralen/UV treatment of HIV-1-infected cells does not cause a significant decrease in the reactivity of HIV-1 core and envelope antigens or cellular antigens to monoclonal antibodies. Experiments are presented demonstrating the use of these cells for flow cytometry studies and for cell surface labeling using the lactoperoxidase 125I iodination procedure.

Productive infection and subsequent interaction of CD4-gp120 at the cellular membrane is required for HIV-induced apoptosis of CD4+ T cells

“…The stock of HIV-ILA ~ was inactivated with 4'-aminomethyltrioxsalen (10 lag/ml) (Calbiochem-Behring) in combination with longwavelength UV light (320 to 380 nm) for 12 min to render the viral preparation non-infectious while preserving most of its antigenicity and ability to bind to CD4 + T cells (Hanson, 1992: Redfield et al, 1981Watson et al, 1990). UV-activated intercalating compunds such as 4/-anainomethyltrioxsalen form covalent adducts with nucleic acids, thereby inactivating cell-free and cell-associated viral infectivity (Redfield et al, 1981 ;Dodd.…”

“…To prepare noninfectious virus, HIV-1LA ~ was treated with 4'-aminomethyltrioxsalen (10 pg/ml) in combination with longwavelength UV light (Watson et al, 1990). This rendered a high titre stock (5 × 107 TCIDs0/ml) essentially noninfectious (< 10 TCIDs0/ml).…”

Human Immunodeficiency Virus Type 1 in Illicit-Drug Solutions Used Intravenously Retains Infectivity

“…In those samples where viral activity in MT-4 cells was not detected at any dilution point after 7 days of incubation, three additional passages were carried out through the addition of fresh MT-4 cells every 3 to 4 days at a ratio of 1:5. A trypan blue exclusion assay was used to monitor cell viability (5). Table 1 shows that HIV-1 was infectious after 20 days of incubation at 22°C in the mixture with the final heroin concentration of 25 mg/ml (the concentration used for injecting by IDUs), and similar results were obtained for HIV-1 mixed with distilled water or RPMI 1640.…”

Bestatin-mediated inhibition of leucine aminopeptidase may hinder HIV infection

“…Syncytium formation was evaluated by light microscopy on days 3, 5 and 7. At various time points, the cells in selected wells were subjected to psoralen/ultraviolet light inactivation (Watson et al, 1990) air-dried and fixed on glass plates with cold acetone. Indirect immunofluorescent staining was performed using a modification of standard procedures (Johnson and Nogueira Araujo, 1981;Aldovini and Walker, 1990).…”