Subcellular distribution of particle-associated antigens in Trypanosoma rhodesiense .

Abstract

Bloodstream forms of the African trypanosome Trypanosoma rhodesiense were subjected to analytic cell fractionation procedures using isopycnic sucrose gradient centrifugation. The fractions obtained were analyzed using fused rocket immunoelectrophoresis, which revealed the most antigenic components to be associated with particles sedimenting between density increments of 1.10 to 1.15 and 1.20 and 1.22. From the distribution of various marker enzymes and the fluorogenic label fluorescamine, these particle populations were identified as derived from flagella pocket and surface membrane, respectively. Crossed immunoelectrophoresis of postulated flagella pocket and surface membrane fractions demonstrated at least seven and 10 distinct antigens, respectively, with only limited cross-reactivity. On the basis of crossed affini-immunoelectrophoresis four to five of each of these membrane antigens were glycoproteins, being precipitated in a concanavalin A-containing gel. Of the various detergent treatments used, it was found that optimum extraction of these glycoprotein antigens was achieved using the zwitterionic detergent Zwittergent 3-12 (0.1%) in the presence of 0.4% Triton X-100. Fused rocket immunoelectrophoresis also revealed a single prominent antigen associated with a particle population having an equilibrium density of approximately 1.180, which was identified as lysosomal from the distribution of marker enzymes. Other intracellular sites were not significantly antigenic.