Figure 3

Anti-Lepp12 antibody responses in patients with clinically and parasitologically diagnosed VL. Reactivities of diagnosis and follow up sera against recombinant Lepp12 from eight VL patients are shown. GST-Lepp12 and GST, purified as described in Methods section, were eluted from gluthatione sepharose with 0.1% SDS in PBS. Recombinant proteins were coated at 0.05 microg per well onto microtiter plate and sequentially incubated with patient serum diluted 1:100 and peroxidase-labeled anti-human IgG. Absorbencies obtained with GST-Lepp12 were corrected for absorbencies measured with GST alone. Sera from leishmanin skin test (LST) negative controls gave OD values below 0.1. The patient's serum with the highest titer at diagnosis was selected for the preparation of the immunopurified anti-Lepp12 antibodies. Typically 20 microg of immunopurified anti-Lepp12 antibody were obtained from 1 ml of serum.