Abstract
The purpose of the current study was to characterize Leishmania cell‐surface antigens by two different methods established for the purification of glycoproteins and proteins, and to point out a useful approach to define their size and mass heterogeneity. L. tropica parasites were initially isolated from patients with active cutaneous leishmaniasis and were then cultured in vitro. The parasite‐cell layer was solubilised with 6 M guanidinium chloride (GuHCl) and subsequently prepared for the purification procedure. The methods used in this work were gel filtration chromatography and isopycnic density‐gradient centrifugation. Because of the presence of a substantial amount of non‐specific proteins in the culture medium, these methods were not effective alone in distinguishing these antigens. However, a good idea of their N‐glycosylated structures could be obtained by using Periodic acid‐Schiffs (PAS) and Con A lectin, and also size and mass heterogeneity. A combination of these methods effected a clear separation of the antigens. Amino acid analysis of the purified antigens was performed to positively identify them as well‐known Leishmania cell‐surface antigen gene products. The results confirmed the presence of more than one cell‐surface antigen on the Leishmania parasite and the combination of gel chromatography and density‐gradient centrifugation could be useful for their isolation.