We have examined the heparin binding proteins from Leishmania (Viannia) braziliensis promastigotes (HBP-Lb) by chromatography assays. The proposed strategy to isolate an enriched fraction of the HBP-Lb consisted of an association of the Triton X-114 method with affinity chromatography in heparin-Sepharose 4B column. SDS-PAGE analysis of the eluted proteins showed two main protein bands (65.0 and 54.5 kDa), while a single protein band was observed in native electrophoresis gel. The hemagglutination property of HBP-Lb over rabbit erythrocytes was confirmed up to 6.3+/-0.5 microg of protein mL(-1). Additionally, we have assayed the potential of HBP-Lb labeled with sulfo-NHS-LC-biotin in binding to nitrocellulose-immobilized gut proteins extracted of Lutzomyia intermedia and Lutzomyia whitmani. The results indicated a similar profile of five ligands (67.0, 62.1, 59.5, 56.0 and 47.5 kDa) in both studied Lutzomyia species. This is the first direct description of this class of protein in L. (V.) braziliensis with a suggestion of its biological activity in the interaction of Leishmania with Lutzomyia gut cells, which maybe a crucial step during this parasite's life cycle.