FIG 3.

The metastable states induced at 0°C show increased sensitivity to antibody neutralization. (A) Effect of brief exposure to cold on sensitivity of viruses containing 89.6 Env to neutralization. Viruses were incubated at 0°C or 37°C in the absence or presence of the indicated inhibitors (MAbs or polyclonal pooled sera from HIV-infected individuals) for 2 h and then added to the cells to measure residual infectivity. Infection is presented as the fold change relative to untreated virus. The green circles represent the calculated synergistic effect between cold and inhibitor (equation 1). The concentrations of all inhibitors used in neutralization assays are provided in Materials and Methods. (B) Reversibility of the effect of cold on HIV-1 neutralization by PS(A) and MAb 447-52D. Viruses were incubated for 1 h with antibody at 0°C, 25°C, or 25°C after 1-h exposure to 0°C. Samples were then added to CD4⁺ CCR5⁺ cells and further cultured at 37°C for 3 days to measure infectivity. (C) Effect of brief (2-h) exposure to 0°C on neutralization sensitivity of viruses containing 89.6(225I, 305E) Env, as shown in panel A. (D) Neutralization of virus containing the cold-resistant 89.6(225I, 305E) Env increases with incubation time at 0°C. Viruses were incubated with MAb 17b, MAb 447-52D, or no antibody at 0°C for the indicated times and then added to cells to measure residual infectivity.