Figure 2. Purification and specificity of LipL32 antibodies from leptospirosis patient sera.

(A) Affinity purification of LipL32-specific antibodies. Recombinant LipL32 [17] was coupled to an AminoLink Plus column. Pooled convalescent sera from 23 individuals with laboratory-confirmed leptospirosis was added to the LipL32-affinity column. The chromatography products were analyzed by gel electrophoresis (Bis-Tris 4–12% NuPage gel, Novex), and Coomassie G250 staining. Abbreviations: LeptoPS, leptospirosis patient sera (pooled); FT, flow-through fraction; W, fraction after washing with PBS; E1-E4, eluted IgG fractions. (B) Extract of 1×10⁸ leptospires (lane WC) or 0.5 µg of recombinant LipL32 (lane rLipL32) were separated by gel electrophoresis, blotted onto PVDF membrane, and probed with affinity purified LipL32 IgG fraction E2 (1∶200).