Validation of a rat pheochromocytoma (PC12)-based cell survival assay for determining biological potency of recombinant human nerve growth factor
| Autor: | Herman Chu, Venkat R. Mukku, Anthony Chen, Barbara Casto, Hélène Gazzano-Santoro, Claire Kotts, Ellen Gilkerson, Eleanor Canova-Davis |
|---|---|
| Rok vydání: | 1999 |
| Předmět: |
Cell Survival
Clinical Biochemistry Pharmaceutical Science Oxazines Pheochromocytoma PC12 Cells Sensitivity and Specificity Fluorescence Analytical Chemistry law.invention law Nerve Growth Factor Drug Discovery Animals Humans Bioassay Potency Fluorometry Coloring Agents Spectroscopy chemistry.chemical_classification Chromatography Dose-Response Relationship Drug Chemistry Reproducibility of Results Biological activity Repeatability Recombinant Proteins In vitro Rats Nerve growth factor Xanthenes Recombinant DNA Biological Assay |
| Zdroj: | Journal of Pharmaceutical and Biomedical Analysis. 21:945-959 |
| ISSN: | 0731-7085 |
| DOI: | 10.1016/s0731-7085(99)00225-3 |
| Popis: | A method has been validated, according to the Guidelines of the International Conference on Harmonization (ICH), for precise quantitation of the biological activity of recombinant human nerve growth factor (rhNGF) for lot release testing. The assay is based on the survival of a subclone of rat pheochromocytoma PC12 cells (PC12-CF) in response to rhNGF. Cell survival is measured by monitoring the reduction, by living cells, of the alamarBlue dye into a red form which is highly fluorescent. The assay is simple, has high throughput (performed in 96-well microtiter plates) and shows reproducible dose-response curves in the concentration range of 0.2-50 ng/ml. The method was validated for its linearity, accuracy, precision, robustness, and to meet current regulatory requirements. The assay demonstrated good linearity, yielding a coefficient of determination of 0.9902. Sample recovery studies demonstrated an accuracy ranging from 96 to 98%. The repeatability of the assay and intermediate precision had coefficients of variation (CV) of |
| Databáze: | OpenAIRE |
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