RESUMO
Serological tests to detect Trypanosoma cruzi antibodies have been used for screening blood donors, for epidemic studies, and for diagnosis of probably infected persons. Among different tests, the enzyme-linked immunosorbent assay (ELISA) with total, semipurified, or synthetic antigens has been widely used, mainly due to its easy automation. Aiming to improve serological studies concerning Chagas' disease, we have developed and evaluated a new test, the TcF-ELISA, using an artificially engineered recombinant antigen, which contains tandem sequences of different T. cruzi-specific peptides. The sensibility of the TcF-ELISA was determined with 101 serum samples from chagasic patients well-defined by clinical and epidemiological criteria. The specificity was determined with 39 serum samples from leishmaniasis or kala-azar patients and 150 serum samples from nonchagasic blood donors from Sao Paulo, Brazil. The TcF-ELISA showed 100% sensitivity and 98.94% of specificity. Compared with conventional ELISA (with semipurified T. cruzi epimastigote antigens), the TcF-ELISA showed advantages; for example, it distinguishes better between reagent and nonreagent serum and provides better precision and a lower occurrence of leishmaniasis cross-reactions. Our studies demonstrate high reproducibility between two different lots of the TcF ELISA and its applicability for the serological diagnosis of Chagas' disease.
Assuntos
Anticorpos Antiprotozoários/sangue , Antígenos de Protozoários/imunologia , Doença de Chagas/diagnóstico , Trypanosoma cruzi/imunologia , Animais , Antígenos de Protozoários/genética , Doença de Chagas/parasitologia , Ensaio de Imunoadsorção Enzimática/métodos , Humanos , Peptídeos/genética , Peptídeos/imunologia , Proteínas Recombinantes/imunologia , Proteínas Recombinantes/metabolismo , Reprodutibilidade dos Testes , Sensibilidade e EspecificidadeRESUMO
No diagnostico sorologico da doenca de Chagas podem ser encontradas acentuadas divergencias mesmo entre laboratorios de grande experiencia. Para a padronizacao de um teste imunoenzimatico destinado primariamente a selecao de doadores de sangue foram cuidadosamente escolhidos paineis de soros que se buscaram como representativos das populacoes de chagasicos e de nao chagasicos. Produzido para maxima sensibilidade e estabilidade, o novo reagente (bioElisa - cruzi) foi testado em 1648 soros, com diagnostico clinico de doenca de Chagas em 219 e de outras afeccoes em 104. O teste foi comparado com testes ja bem padronizados, de imunofluorescencia (IF) e de hemaglutinacao (HA), em 1325 doros. O limiar de reatividade (cut off), estabelecido como ideal, foi indicado nos testes pela absorbancia de um soro de reatividade minima. A sensibilidade do teste imunoenzimatico foi de 0,9954 e a especificidade, como conegatividade, de 0,9969. Nao foram vistos resultados falso-positivos em casos de sifilis, toxoplasmose, mononucleose e de soros com altos titulos de anti-estreptolisina 0, mas foram encontrados em 5 de 15 casos de leishmaniose tegumentar, 10 de 24 casos de calazar, 1 de 15 casos de artrite reumatoide e 1 de 12 casos de lupus eritematoso sistemico....
Assuntos
Animais , Humanos , Doadores de Sangue , Doença de Chagas/diagnóstico , Técnicas Imunoenzimáticas/normas , Valor Preditivo dos Testes , TriagemRESUMO
In the serological diagnosis of Chagas disease large divergences may be found even between laboratories with experience, as a consequence of different criteria for the standardization of the tests. To standardize a immunoenzymatic test developed primarily for screening blood donors, serum panels were carefully chosen so as to best represent chagasic and non-chagasic populations. Produced for the highest sensibility and stability, the new reagent (bioELISA cruzi, Biolab Diagnóstica S/A, Brasil), was tested in serum from 1648 patients 219 with Chagas disease and 104 with other diseases, plus a comparison with well standardized immunofluorescence and hemagglutination tests in 1325 sera. In the immunoenzymatic assays, the cut off was indicated by the absorbance value of a chagasic serum showing a minimal reactivity. ELISA sensibility was 0.9954 and specificity 0.9969, as co-negativity. False positive results were absent with sera from syphilis, toxoplasmosis, mononucleosis and high titered sera for antistreptolysin 0 antibodies. However they were seen in 5 to 15 cases of tegumentar leishmaniasis, 1 of 12 Kala-azar 1 of 15 rheumatoid arthritis and 1 of 12 systemic lupus erythematosus. The high sensibility in chagasics and high specificity in the general population indicate the confiability of the immunoenzymatic assay for screening blood donors and even to confirm a clinical diagnosis of Chagas' disease.