Abstract In addition to generating antibodies against infections, B lymphocytes produce cytokines to regulate immune functions. Here, we have identified B cells as a major source of cytokine IL-27 (the heterodimer of IL-27p28 and EBI3) by using B cell-deficient mMT and JH mice, mice with B cell-specific deficiency in Ebi3 and NSG mice transplanted with highly purified B cells. IL-27 in turn targets B cells to undergo class-switching to IgG2a, as shown by decreased antigen-specific IgG2a in mice with B cell-specific deficiency IL-27 receptor signaling, leading to defective clearance of vaccinia virus infection. The paracrinic function of IL-27 is recapitulated in vitro by B cell production of IL-27 upon stimulation by a TLR ligand together with CD154 (CD40 ligand) and IL-21, two hallmark stimuli of TFH cells, and IL-27 induction of T-bet and a gene signature similar to that induced by IFNγ, IL-27 collaboration with IFNγ in inducing T-bet and IgG2a, and, finally, selective switching to IgG2a in primed B cells when mixed with IL-27-producing B cells, which display a metabolic program distinct from that in IL-27-responding B cells, i.e., heightened TCA cycle activities and enhanced lactate generation and glutaminolysis. B cell-produced IL-27 also promotes differentiation of TFH cells, as shown by reduced number of TFH cells and defective IL-21 expression in residual TFH cells in mice with B cell-specific deficiency in Ebi3 and, consequently, severe impairments in the germinal center development, class-switching to all Ig isotypes and protective antibody responses to vaccinia virus infection in these mice. Overall, B cell production of IL-27 is a previously unrecognized important regulatory mechanism in the antibody response and anti-viral immunity.