RESUMO
Streptokinase is widely used as an anticoagulant drug for the treatment of heart attacks. Because of antibody production against injected drug in individuals consuming streptokinase and causing allergic reactions, streptokinase treatment effects become neutral. Recombinant mutant type of streptokinase was prepared by removing of 42 amino acids from the C terminal region. ELISA plates were coated by natural and mutant streptokinase as antigen. Ninety-six normal serum samples as well as 27 streptokinase consumer serum samples (patients with acute myocardial infraction) were analyzed. The results showed that serum antibodies against natural streptokinase were three times more than those against the mutated streptokinase. In case of preserving thrombolytic activity, mutated streptokinase can be used as an alternative of the natural form.
Assuntos
Anticorpos Antibacterianos/sangue , Anticoagulantes/metabolismo , Anticoagulantes/uso terapêutico , Infarto do Miocárdio/tratamento farmacológico , Estreptoquinase/metabolismo , Estreptoquinase/uso terapêutico , Terapia Trombolítica , Sequência de Aminoácidos , Anticorpos Antibacterianos/biossíntese , Anticorpos Antibacterianos/imunologia , Anticoagulantes/imunologia , Ensaio de Imunoadsorção Enzimática , Humanos , Dados de Sequência Molecular , Infarto do Miocárdio/sangue , Infarto do Miocárdio/imunologia , Estrutura Terciária de Proteína , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/uso terapêutico , Deleção de Sequência , Estreptoquinase/genética , Estreptoquinase/imunologiaRESUMO
BACKGROUND: Although a simple and direct method does not exist for the detection of chlamydial infections, there are situations in which reliable serological tests, with sensitivity related to a specific antigen, could be helpful. OBJECTIVE: The aim of this study was to clone the first 1100 bp of the C. trachomatis outer membrane protein 2 (omp2) gene in order to prepare a recombinant protein for use in an ELISA system designed to recognize the anti- C. trachomatis antibody in patient sera. METHODS: The PCR product of the chlamydial omp2 gene was cloned in pBluescript and its first 1100 bp was subcloned in the pQE-30 expression vector and induced by IPTG. The recombinant protein was purified by affinity chromatography and its purity was confirmed by SDS-PAGE, gel diffusion and western blot analyses. The purified protein was coated onto a polystyrene microplate and tested by ELISA using patient serum. RESULTS: We have cloned, over-expressed and purified biologically functional recombinant truncated Omp2 from C. trachomatis for use, as a species-specific recognition antigen, in an ELISA system. In this study we determined a cut-off value of 0.345 for this ELISA system using 55 negative sera and measured six positive sera at dilutions of 1:20-1:2560. CONCLUSION: As a species-specific recognition antigen, the over-expressed and purified recombinant truncated Omp2 from C. trachomatis performed well in an ELISA system.
