Lipoteichoic acid (LTA) of Gram-positive bacteria is regarded as the counterpart biomolecule of lipopolysaccharide (LPS) of Gram-negative bacteria because of their structural and immunological similarities. Although LPS induces a strong polyclonal expansion of B cells, little is known about the effect of LTA on B cell proliferation. In the present study, we prepared LTAs from Gram-positive bacteria and examined their effect on splenic B cell proliferation. Unlike LPS, LTA did not induce B cell proliferation. Instead, Staphylococcus aureus LTA (Sa.LTA) appeared to inhibit LPS-induced B cell proliferation in vitro, ex vivo, and in vivo models. Such effect was observed neither in splenocytes from Toll-like receptor 2 (TLR2)-deficient mice nor in the purified splenic B cells. Furthermore, decreased ERK phosphorylation appeared to be responsible for this phenomenon. Collectively, our results support that Sa.LTA inhibited LPS-induced B cell proliferation through the decrease of ERK phosphorylation via TLR2 signaling pathway.