Identification of specific and cross-reactive antigens of Leishmania donovani chagasi by human infection sera

Cloned Leishmania donovani chagasi (Ldc) promastigotes were analyzed by SDS-PAGE separation and immunoblotting with human infection sera. The patterns of antigen reactivity were compared by using sera from individuals with Ldc, Leishmania mexicana amazonensis (Lma), Trypanosoma cruzi, Mycobacterium tuberculosis, or Mycobacterium leprae infections. Sera from individuals with these infections recognized Ldc antigens in several m.w. ranges. Reactivity was due to recognition of Ldc molecules and not to Ldc culture medium components, as shown by comparing Ldc promastigotes grown in the presence or absence of fetal bovine serum (FBS), by immunoblotting of FBS, and by [35S]methionine labeling. The major findings of the study were as follows. Immunoblots with Ldc promastigotes could be used to distinguish individuals with Ldc infections from those with Lma infections. Persons with Ldc infections had antibodies to a Ldc antigen of approximately 32 to 35 kd not recognized by persons with Lma infections. Individuals cured of acute Ldc infection did not develop antibodies that differed in specificity to those present during their acute phase of infection. Ldc antigens in the 62 to 66 kd region were recognized by all individuals with Ldc or Lma infections but were not recognized by individuals in the other disease groups or by control sera. This region was found to contain at least four distinct bands, one of which appeared to be glycosylated as indicated by periodic acid-Schiff staining and concanavalin A labeling; an apparently nonglycosylated protein of 62 to 63 kd was eluted from SDS-PAGE gels and was used to diagnose Ldc infection by the ELISA. Whereas crude Ldc antigen gave false positive results with T. cruzi and mycobacteria infection sera, the eluted 62 to 63 kd protein was 100% specific and sensitive in the diagnosis of Ldc infection.