pTFH cells promote B-cell Ab production and differentiation on H1N1-specific stimulation. Cryopreserved PBMCs obtained 4 weeks after H1N1/09 vaccination were thawed and rested overnight. B cells (CD20⁺) were isolated with magnetic beads and pTFH (CD3⁺CD4⁺CD45RA⁻CXCR5⁺) and non-pTFH (CD3⁺CD4⁺CD45RA⁻CXCR5⁻) cells were purified by cell sorting. B cells were cocultured with either pTFH or non-pTFH cells at a 1:1 ratio in medium alone or in the presence of 5 μg/mL of H1N1 vaccine Ag or 1 μg/mL of staphylococcal enterotoxin B for 7 days. (A,C) Culture supernatants from the pTFH + B-cell cocultures (A) or non-pTFH + B-cell cocultures (C) were harvested and IgG production was measured by ELISA. (B,D) Cells were harvested and frequency of plasmablasts (defined as CD20^lowCD21^low/−CD27⁺CD10⁻Ki67⁺ events) within the B cells was evaluated by flow cytometry in the pTFH + B-cell cocultures (B) or the non-pTFH + B-cell cocultures (D). Bars represent the means and error bars indicate the SD of 3 HCs, 3 HIV⁺ H1N1/09 vaccine responders (R), and 3 HIV⁺ vaccine nonresponders (NR).