Impact of B-cell depletion after rituximab administration on anti-SIVsmm humoral immune responses. (A) ELISA testing of anti-SIVsmm gp41 antibodies showed significant delay in the seroconversion patterns in RMs with complete tissue CD20⁺ B-cell depletion (red symbols and lines) and no significant difference between control RMs (black symbols and lines) and rituximab-treated RMs with incomplete tissue CD20⁺ B-cell depletion (green symbols and lines). (B) Western blot testing on serial samples confirmed the trend observed by ELISA, with no significant difference in the seroconversion patterns between control RMs (illustrated by RMs DG34 and DH44) and rituximab-infused RMs with incomplete tissue depletion (illustrated by RMs DG04 and CA16). Significant delays in the WB seroconversion were observed in RMs with complete CD20⁺ B-cell depletion after rituximab infusion (illustrated by RMs CF03 and BT49). Neutralizing antibody testing against both highly susceptible TCLA-SIVmac251 strain (M7-Luc assay; C) and SIVsmmD215 strain (TZM-bl assay; D) showed delayed seroconversion and lower titers in RMs infused with rituximab. RMs showing complete tissue CD20⁺ B-cell depletion showed the lowest Nab titers. Day 0 corresponds to SIV inoculation. Rituximab was infused every 21 days beginning 1 week before SIV inoculation. RMs DG04, CA16, CF03, and BT49 received 4 rituximab administrations. RMs EI74, EI09, and EL55 received rituximab for up to 160 days pi.