Immunological characteristics and localization of the Trichinella spiralis glutathione S-transferase.
Trichinella spiralis glutathione S-transferase (TsGST) was isolated from crude extracts of L1 larvae by glutathione-affinity chromatography. Two closely migrating polypeptides with molecular masses of 28.5 and 28 kDa were identified by electrophoresis. Three isoforms of pI 5.6, 5.8, and 6.0 were detected by isoelectric focusing. Purified TsGST showed a low transferase activity as measured with 1-chloro-2,4-dinitrobenzene; glutathione peroxidase activity was also demonstrated using cumene hydroperoxide. A rabbit antiserum against TsGST reacted by western blot with crude extracts of Trichinella britovi and Trichinella nativa but not with extracts of Trichinella pseudospiralis, Fasciola hepatica, Schistosoma bovis, Schistosoma mansoni, Dirofilaria immitis, Toxocara canis, or Anisakis sp. TsGST was detected by western blot in extracts of T. spiralis adults, but not in newborn larvae or L1 excretory-secretory products; yet, an antiserum against T. spiralis excretory-secretory products reacted with TsGST. By immunoelectron microscopy, TsGST was found in the granules of the alpha- and beta-stichocytes of L1 larvae, as well as in some granules of the stichocytes of 72-hr adults. Rabbits experimentally infected with T. spiralis developed substantial levels of anti-TsGST antibodies. Moreover, circulating TsGST was detected in serum by a sandwich-enzyme-linked immunosorbent assay, isolated from serum by glutathione-affinity chromatography, and characterized as TsGST by western blot.
Rojas
J
J
Departamento de Immunologia Biología Celular, Instituto de Parasitologia y Biomedicina López-Neyra, CSIC, Granada, España.
Rodríguez-Osorio
M
M
Gómez-García
V
V
D000818:Animals; D000909:Antibodies, Helminth / Q000096:biosynthesis*; D015153:Blotting, Western; D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes; D005978:Glutathione / Q000378:metabolism; D005979:Glutathione Peroxidase / Q000032:analysis; D005982:Glutathione Transferase / Q000032:analysis* / Q000276:immunology* / Q000302:isolation & purification; D007106:Immune Sera / Q000276:immunology; D007525:Isoelectric Focusing; D007814:Larva / Q000201:enzymology / Q000276:immunology; D016253:Microscopy, Immunoelectron; D008970:Molecular Weight; D011817:Rabbits; D017160:Trichinella spiralis / Q000201:enzymology* / Q000276:immunology / Q000648:ultrastructure; D014235:Trichinellosis / Q000201:enzymology / Q000276:immunology / Q000469:parasitology*
D000909:Antibodies, Helminth; D000939:Epitopes; D007106:Immune Sera; D005979:Glutathione Peroxidase; D005982:Glutathione Transferase; D005978:Glutathione