A simple immune complex dissociation ELISA for leishmaniasis: Standardization of the assay in experimental models and preliminary results in canine and human samples

摘要

Visceral leishmaniasis, caused by Leishmania (Leishmania) chagasi, is a chronic parasitic disease of humans and dogs. Confirmation of the protozoal agent in bone marrow, lymph node or spleen aspirate is diagnostic, while specific-IgG serology is used mainly for epidemiology despite the general presence of high levels of serum immunoglobulin. Anecdotal reports of false-negative serology in active disease cases are known and are ascribed to the formation of immune complexes. Because dissociation of immune complexes can be accomplished by acid treatment, we devised a simple, routine enzyme immunoassay (ELISA) for the dissociation of immune complexes in serum samples using acid treatment in wells adsorbed with Leishmania antigen (dELISA). Confirmatory acid dot-blot was also developed for antigen detection by anti-Leishmania rabbit antiserum. In experimental L. chagasi hamster models, immune complexes interfered with ELISA mostly in the 30 and 60 days postinfection, according to both dELISA and antigen dot-blot results. In larger samples from endemic areas, dELISA was positive in 10% of seronegative dog samples (7/70) and 3.5% in negative human samples (3/88), showing that dELISA could be used in the serodiagnosis of visceral leishmaniasis. Moreover, dELISA could be used as an alternative approach to screening asymptomatic visceral leishmaniasis patients, instead of invasive confirmatory testing.