RESUMO
The goal of this work was to study sensitivity and specificity of the developed ELISA set for the identification of IgG antibodies against Chlamydia trachomatis HSP-60 (using biotinylated tyramine-based signal amplification system). The study was conducted using a panel of characterized sera, as well as two reference ELISA sets of similar purpose. According to the results of ELISA informative value parameters, the ELISA we have developed showed the highest specificity and sensitivity parameters (no false negative or false positive results were registered). In 4 out of 15 intralaboratory panel serum samples initially identified as negative, anti-HSP-60 IgG-antibodies test result in reference ELISA sets upon dilution changed from negative to positive. The nature of titration curves of false negative sera and commercial monoclonal antibodies Ð57-Ð9 against C. trachomatis HSP-60 after incubation for 24 h was indicative of the presence of anti-idiotypic antibodies in these samples. Upon sera dilution, idiotypic-anti-idiotypic complexes dissociated, which caused the change of test result. High informative value of the developed ELISA set for identification of IgG antibodies against C. trachomatis HSP-60 has been proven. Anti-idiotypic antibodies possessing C. trachomatis anti-HSP-60 activity and being one of the causes of false negative results of the relevant ELISA-based tests have been identified in blood sera of individuals infected with chlamydial genitourinary infection agents.
Assuntos
Anticorpos Antibacterianos/análise , Antígenos de Bactérias/sangue , Chaperonina 60/sangue , Infecções por Chlamydia/diagnóstico , Chlamydia trachomatis/imunologia , Ensaio de Imunoadsorção Enzimática/métodos , Imunoglobulina G/análise , Anticorpos Anti-Idiotípicos/química , Anticorpos Antibacterianos/sangue , Anticorpos Antibacterianos/imunologia , Complexo Antígeno-Anticorpo/química , Antígenos de Bactérias/imunologia , Biotinilação , Chaperonina 60/imunologia , Infecções por Chlamydia/sangue , Infecções por Chlamydia/imunologia , Infecções por Chlamydia/microbiologia , Chlamydia trachomatis/química , Ensaio de Imunoadsorção Enzimática/normas , Reações Falso-Negativas , Humanos , Soros Imunes/química , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Sensibilidade e Especificidade , Tiramina/químicaRESUMO
Validation procedure for method of monitoring the biological activity of reÑombinant human interleukin-7 has been developed and conducted according to the requirements of national and international recommendations. This method is based on the ability of recombinant human interleukin-7 to induce proliferation of T lymphocytes. It has been shown that to control the biological activity of recombinant human interleukin-7 peripheral blood mononuclear cells (PBMCs) derived from blood or cell lines can be used. Validation characteristics that should be determined depend on the method, type of product or object test/measurement and biological test systems used in research. The validation procedure for the method of control of biological activity of recombinant human interleukin-7 in peripheral blood mononuclear cells showed satisfactory results on all parameters tested such as specificity, accuracy, precision and linearity.