Molecular and CellularBiochemistry 109: 25-36, 1992. © 1992 KluwerAcademic Publishers. Printedin the Netherlands. Trypanosoma cruzi glycoprotein 72: Immunological analysis and cellular localization Guenter Harth 1'2, Alea A. Mills 1, Thais Souto-Padr6n 3 and Wanderley de Souza 3 Department of Immunology and Infectious Diseases, Research Institute, Palo Alto Medical Foundation, Palo Alto, California 94301, and 2Department of Medicine, Stanford University School of Medicine, Stanford, California 94305 and 3Laborat6rio de Ultraestrutura Celular e Microscopia EletrOnica, Instituto de Biofisica Carlos Chagas Filho, Universidade Federal do Rio de Janeiro, Ilha do Fundao, 21941, Rio de Janeiro, Brazil Received30 April 1991; accepted 19 July 1991 Abstract Two monoclonal antibodies were used to biochemically characterize glycoprotein 72 (GP72) from Trypanosoma cruzi and to localize the protein in live and fixed parasites by indirect immunofluorescence and in thin section of parasites by immunogold electron microscopy. GP72 was shown in immunoblots to be specific for the epimastigote stage; the protein could not be detected in trypomastigotes. Each antibody reacted with a different epitope on the glycoprotein and deglycosylation of GP72 ablated reactivity with one of the antibodies. Indirect immunofluorescence and electron microscopic evaluation of parasite associated gold particles showed the presence of GP72 in the cell surface membrane including the flagellar pocket and the cytostome. In addition, cytoplasmic membrane vesicles of the endosomal- lysosomal system stained intensely. (Mol Cell Biochem 109: 25-36, 1992) Key words: T. cruzi, glycoprotein 72 (GP72), stage specificity, electron microscopy Introduction The protozoan parasite Trypanosoma cruzi is the causa- tive agent of Chagas' disease, a chronic disease afflict- ing an estimated 10-20 million people in Latin America [1]. As the parasite passes between its two hosts, reduvi- id insects and mammals, it undergoes a complex series of morphological and physiological conversions. The epimastigote and amastigote stages are the multiplica- tive forms found in the insect vector and within mam- malian host cells, respectively. The trypomastigote stage represents the highly infectious, nonreplicative form found as bloodform trypomastigotes in the blood- stream of infected mammalian hosts and in the posteri- or portion of the intestine of the invertebrate host as metacyclic trypomastigotes [2]. Surface membrane proteins of T. cruzi have been studied by immunological methods, lectin binding, and two dimensional gel electrophoresis [3-6]. These stud- ies have led to the identification of stage-specific surface components and confirmed earlier results that demon- strated many surface components to be glycoproteins Address for offprints: G. Harth, Department of Immunologyand InfectiousDiseases, Research Institute, Palo Alto MedicalFoundation, Palo Alto, California 94301, USA