FIG 6.

Immunocapture of NL4.3-derived Del 102-114 mutant virus and NL4.3-derived virus carrying Del 505-514 mutant ICAM-1. (A) NL4.3-derived WT and mutant virus particles were prepared in 293T cells stably expressing wild-type ICAM-1. (B) NL4.3-derived WT viruses were produced in 293T cells transiently expressing either wild-type ICAM-1 or Del 505-514 mutant ICAM-1. Next, virus stocks were captured by using magnetic beads coated with an anti-ICAM-1 antibody (R6.5) or an isotype-matched irrelevant biotinylated antibody (used as a control). Precipitated viral entities were quantified with a homemade p24 ELISA. Data shown represent the means ± standard deviations of data from triplicate samples (i.e., total amounts of virus captured with anti-ICAM-1 divided by total amounts of virus captured with the control antibody) and are representative of three independent experiments. (C and D) Flow cytometry analyses of ICAM-1 surface expression in 293T cells either stably (C) or transiently (D) expressing ICAM-1. Wild-type ICAM-1 is depicted as a solid black line (C and D), Del 505-514 mutant ICAM-1 is shown as a solid gray line (D), and the control is presented as a gray-filled histogram. Data shown represent cell surface ICAM-1 expression after 1 month of selective drug pressure for stable transfectants (C) and 2 days after transfection for transient transfectants (D). Statistical analyses were made by using the Student t test, and asterisks denote a statistically significant difference between the NL4.3 WT or NL4.3 WT CD54 WT and the listed mutants (****, P < 0.0001).