RESUMO
PURPOSE: The implementation of an in vivo assay to determine the biological activity of human recombinant erythropoietin (Hu-r EPO) is essential. The purpose of this study was to perform and optimize the conditions of an easy in vivo bioassay suitable for routine testing of quality control of Hu-r EPO preparations. MATERIAL AND METHODS: Normocythemic 8 weeks female mice treated with different Hu-r EPO doses were employed. The reticulocyte response was measured by flow cytometry and by visual count in a Neubauer cell count chamber, after selective red blood cell haemolysis. A unique subcutaneous injection with blood extraction 96 hours later was the schedule employed. The reticulocyte count measured by both methods was plotted against the log dose of Hu-r EPO. RESULTS: The dose-response curve obtained was linear between 5 and 160 UI/mouse and the doses chosen for future assays were 10, 30 and 90 UI/mouse. The use of at least 6 animals per dose and not less than 3 assays to obtain reliable limits according to international regulations is convenient. Thirty assays were performed in four different samples and were analyzed by parallel lines (3 + 3) relating the response with the log dose. The coefficient of correlation between both methods was 0.989, so they are equivalent. CONCLUSIONS: This method is suitable because fewer animals and bioassays are necessary to obtain fiducial limits according to international requirements. It is in agreement with the tendency to reduce the number of animals used for bioassay because ethical and economic reasons.
Assuntos
Bioensaio/métodos , Contagem de Eritrócitos/efeitos dos fármacos , Eritropoetina/farmacologia , Contagem de Reticulócitos/efeitos dos fármacos , Animais , Bioensaio/economia , Separação Celular , Relação Dose-Resposta a Droga , Eritropoetina/administração & dosagem , Eritropoetina/normas , Estudos de Avaliação como Assunto , Feminino , Citometria de Fluxo , Humanos , Injeções Subcutâneas , Camundongos , Controle de Qualidade , Proteínas Recombinantes , Reprodutibilidade dos Testes , Sensibilidade e EspecificidadeRESUMO
A study was carried out to establish an appropriate method for streptokinase (SK) potency determination (biological assay) in order to fulfil the main function of the Instituto Nacional de Medicamentos respecting products marketed in Argentina. The potency of different commercial samples of SK was determined against the International Standard, and three internationally accepted methods were used for this purpose: fibrin plate, clot lysis and chromogenic method. The analysis of results suggests that the fibrin plate method is the least precise and reproducible. The clot lysis and chromogenic methods demonstrated great precision and reproducibility, giving a correlation coefficient of 0.99. It is concluded that both of these methods are best suited to determine potency of SK commercial products.
Assuntos
Fibrinolíticos/farmacologia , Estreptoquinase/farmacologia , Animais , Bovinos , Compostos Cromogênicos/metabolismo , Estudos de Avaliação como Assunto , Fibrina/metabolismo , Fibrinólise/efeitos dos fármacos , Fibrinolíticos/análise , Fibrinolíticos/normas , Oligopeptídeos/metabolismo , Padrões de Referência , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Estreptoquinase/análise , Estreptoquinase/normasAssuntos
Heparina/normas , Academias e Institutos , Animais , Argentina , Padrões de Referência , SuínosRESUMO
In agreement with the Regional Programme of Reference Materials of the Panamerican Health Organization the Instituto Nacional de Farmacología y Bromatología of Buenos Aires designed a study for the calibration of a Reference Material for Heparin, porcine, mucosal and a Reference Material for Heparin, bovine, mucosal. The assay methods used in this study were those described in the United States Pharmacopeia XXI Ed and British Pharmacopoeia 1980, Addendum 1983. The overall combined potency estimates of both heparin in preparations relative to 4th Int.St. was 1633.83 UI/ampoule (95% confidence limits 1609.70-1657.96 UI/ampoule) for porcine heparin and 1332.31 UI/ampoule (95% confidence limits, 1302.31-1361.77 UI/ampoule) for bovine heparin. The assigned unitage was 1630 UI/ampoule for the porcine Reference Material and 1330 UI/ampoule for the bovine Reference Material.