Clone m17 4

To assess the importance of LFA-1, CTLs were pre-incubated with LFA-1 blocking antibody (20 µg/mL, Clone M17/4, BioXCell) or an IgG2aκ isotype control antibody (20 µg/mL, Clone RTK2758, BioLegend) at 37 °C for 5 min before the addition of target cells/stimulatory beads. The final concentration of antibodies during the assay was 10 µg/mL. Human LFA-1 was blocked using the anti-CD18 clone TS1/18 (20 µg/mL, Invitrogen), with a mouse IgG2a antibody (Sigma) serving as an isotype control. To induce T-cell activation independently of the TCR, CTLs were pre-incubated with phorbol myristate acetate (PMA, 20 ng/mL, Sigma Aldrich) and the Ca²⁺ ionophore A23187 (2 µM, Tocris Bioscience) at 37 °C for 5 min before the addition of target cells/stimulatory beads, yielding final concentrations of 10 ng/mL PMA and 1 µM A23187. Additional information about antibodies may be found in Supplementary Table 1.

Chronically infected mice (4 weeks p.i.) were treated on days 1 and 3 of the treatment regimen with 200 μg i.p. each of anti-LFA-1 (Bio X Cell, clone M17/4, cat# BE0006) and anti-VLA-4 (Bio X Cell, clone PS/2, cat# BE0071) blocking antibodies or control IgG. They were then sacrificed and analyzed on day 5.