Trypanosoma cruzi: quantification in tissues of experimentally infected mice by limiting dilution analysis.

A limiting dilution assay (LDA) was developed for the quantification of Trypanosoma cruzi in the heart and blood of infected mice. Three groups of swiss mice were injected ip with "CL", "Colombiana," and "Y" strains. At 1-day intervals after infection, blood and the heart were removed. Serial blood dilutions in LIT medium were performed and distributed in four groups of 24 microplate wells. The growth of parasite was visually checked in an inverted microscope. It was found that curves of parasitemia obtained by parasite counting in a hemocytometer or estimated by LDA were similar. A similar method was used to quantify parasites in the heart of mice. The heart was cut, washed, dried, and its weight was determined. The heart pieces were disrupted by passage through a mesh stainless-steel screen into LIT. Serial dilutions of the heart homogenate were made in LIT and added to at least 24 replicate microplate wells. Parasites were detectable earlier in the heart of mouse infected with Y strain when compared to CL and Colombiana strains. Parasites were detected in the heart of mice of all strains by 6 days after infection. This LDA for quantification of T. cruzi permits a more precise evaluation of the number of living parasites in infected tissues.

Functional analysis of the murine T lymphocyte immune response to a protozoan parasite, Leishmania tropica.

Os resultados apresentados nesta revisao, sumariam uma serie de experimentos planejados no sentido de caracterizar a resposta imune de linfocitos T de camundongos, para o protozoario parasita Leishmania tropica. Populacoes enriquecidas de linfocitos T e clones de linfocitos T especificos para antigenos de L. tropica foram derivados de ganglios linfaticos de camundongos primados e a seguir mantidos em cultura continua in vivo. Ficou demonstrado que estes linfocitos T eram capazes de: A) Expressar o fenotipo de superficie celular Lyt 1+ 2-, B) Proliferar en vitro especificamente em resposta aos antigenos parasitarios quando em presenca de macrofagos singenicos irradiados, C) Transferir uma resposta tipo hipersensibilidade retardada antigeno especifico a camundongos normais singenicos, D) Induzir ativacao especifica de macrofagos parasitizados in vitro resultando em destruicao dos parasitas intracelulares, E) Providenciar uma atividade "helper" especifica para uma resposta com producao de anticorpos in vitro em um sistema haptenocarreador. Estudos de protecao utilizando estas populacoes definidas de linfocitos T poderao permitir a caracterizacao de antigeno(s) parasitarios implicados na inducao de uma resposta imune celular com beneficios para o hospedeiro