Western Blot Analysis of Sera from Leishmania major-Infected BALB/c and C57BL/6 Mice

In Leishmania (L.) major-infected BALB/c mice Th2-type cells results in disease progression, whereas C57BL/6-infected mice mount a Th1-type response, which leads to control of the infection. Th2 response correlates with IgG1 whereas, Th1 response supports switching to IgG2a subclasses. Since IgG isotype-dominated response depends on different CD4 + T cell subsets, we studied the antigenic profile of L. major promastigotes that induce IgG1, IgG2a and IgG2b isotypes in BALB/c and C57BL/6 mice to provide insight into Th1 and Th2 inducing antigens. Humoral immune responses were studied in sera from L. major-infected BALB/c and C57BL/6 mice by ELISA and immunoblot analysis. These techniques were used to detect IgG, IgG1, IgG2a, and IgG2b antibodies against crude promastigote antigens. BALB/c mice showed higher IgG, IgG1 and IgG2a antibody levels compared to C57BL/6 mice (p<0.01). The IgG2a/IgG1 ratio was higher in C57BL/6 mice. There was no significant difference in IgG2b level between susceptible (BALB/c) and resistant (C57BL/6) mice. Western blot analysis revealed that 27, 32, 117, 125 and 153 kDa bands reacted only with IgG1 from both mice strains. The 41, 45, 47, 52 and 55 kDa bands reacted only with IgG2a from C57BL/6. A very similar pattern of immunostaining was observed for IgG2a and IgG2b in each mouse strain. The results from both mouse strains indicate that some antigens react exclusively with IgG1, the antibody, which may be involved in generation of a predominant Th2 response. The antigens that reacted strongly with IgG2a in C57BL/6 are potential candidates for eliciting protective Th1 response.