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The paraflagellar rod proteins (PAR) purified from Trypanosoma cruzi epimastigotes were shown to protect mice against an otherwise lethal challenge inoculum of 10 3 bloodstream-form trypomastigotes. The injection route used for immunization was shown to have a marked impact on the development of protective immunity. Mice receiving subcutaneous (s.c.) injections of PAR proteins had reduced bloodstream parasitemias and showed 100% survival following challenge. In contrast, mice immunized via the intraperitoneal (i.p.) route developed parasitemia levels equivalent to those of unimmunized controls and did not survive infection. Western blotting (immunoblotting) demonstrated that sera from both i.p. and s.c. immunized mice reacted specifically with PAR proteins; however, the antibody titer of the i.p. immunized mice was approximately 64-fold greater than that of the s.c. immunized mice, suggesting that the protective response in the s.c. immunized mice is cell mediated rather than humoral.Trypanosoma cruzi is a parasitic hemoflagellate that causes Chagas' disease, a major human health problem in Central and South America (4). Chemotherapeutic agents have limited effectiveness against the disease, and no vaccines are available for disease prevention. Experimental vaccines consisting of attenuated forms of the parasite, crude parasite lysates, partially purified proteins, and synthetic peptides derived from the sequence of a 24-kDa excretory protein (ESA) have been tested in laboratory animals with varying degrees of success (reviewed in references 2, 5, 7, 16, and 22). Several of these studies show that susceptible hosts may be partially protected against death by vaccination prior to challenge; however, individual antigens capable of ensuring survival against an otherwise lethal challenge have yet to be identified.In an attempt to identify and purify protective antigens, we focused on proteins present in the flagellum of T. cruzi. The rationale for this approach is based on studies demonstrating that vaccination with crude flagellar extracts are effective in both inducing protective immunity (i.e., 90% survival) in mice after challenge (15,20,21) and reducing tissue lesions in immunized mice compared with nonimmunized mice following a challenge infection (14). In contrast, immunization with a membrane-enriched fraction from the body of the parasite was less effective in inducing a protective response, suggesting that the protective antigen(s) may be unique to the flagellum. A potential candidate for such antigens is the paraflagellar rod, a prominent structure present only in the flagellum of the parasite (5). We have shown that the paraflagellar rod contains two distinct proteins, PAR 1 and PAR 2, and have determined conditions that allow these proteins to be isolated in amounts and purity sufficient for immunization trials (3,18).In this paper, we report the ability of the purified PAR proteins to induce an immune response in mice capable of protecting against an otherwise lethal inoculum of T. cruzi trypomastigote...

Adenovirus type 2 mRNA was translated in S30 extracts from Ehrlich ascites and wheat embryo cells. The in vitro products were identified by sodium dodecyl sulfate-gel electrophoresis after immunoprecipitation with specific antisera in the presence of urea. Seven virion polypeptides could be identified by immunoprecipitation. Three of these appear to be precursors to polypeptides of the virion. mRNA isolated late in adenovirus infection was separated into three size classes by zonal sedimentation. Material sedimenting at 26S was translated into polypeptides corresponding to the largest virion polypeptides II to IV, a 22S fraction corresponding to polypeptide V, and smaller polypeptides and a 15S fraction corresponding to polypeptide IX. A significant amount of polypeptide IX was also synthesized by the 26S and 22S RNA.

The synthesis of adenovirus type 2 (Ad2)-induced early polypeptides was examined in vivo and in vitro by a combination of sodium dodecyl sulfate-polyacrylamide gel electrophoresis alone and specific immunoprecipitation followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Analysis of total [35S ]methionine-labeled polypeptides synthesized in vivo at 3 h postinfection allowed us to detect in infected cells at least 13 distinct polypeptides that are either absent or less conspicuous in extracts from mock-infected cells. These Ad2-induced early polypeptides have molecular weights ranging from 72 x 103 to 10.5 x 103 and have accordingly been designated as E72K to E10.5K. Nine of the in vivo synthesized early polypeptides can be precipitated specifically from infected cell extracts by antisera with specificity against early adenovirus proteins. In vitro translation of mRNA extracted from mock-infected cells and from Ad2-infected cells was carried out in preincubated Ehrlich ascites cell extracts. All the early Ad2-induced polypeptides identified in the extracts from infected cells labeled in vivo were also detected among the polypeptides immunoprecipitated specifically from the in vitro reaction mixtures programmed by RNA extracted at 4 h postinfection from Ad2-infected cells. MATERIALS AND METHODS Cells and virus infection. HeLa cells were grown in suspension cultures in Eagle spinner medium supplemented with 7% calf serum and infected with 865

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