ABSTRACT
Background: Existence of bacterial host-cell DNA contamination in biopharmaceutical products is a potential risk factor for patients receiving these drugs. Hence, the quantity of contamination must be controlled under the regulatory standards. Although different methods such as hybridization assays have been employed to determine DNA impurities, these methods are labor intensive and rather expensive. In this study, a rapid real-time PCR test was served as a method of choice to quantify the E. coli host- cell DNA contamination in widely used recombinant streptokinase [rSK], and alpha interferon [IFN-alpha] preparations
Methods: A specific primer pair was designed to amplify a sequence inside the E. coli 16S rRNA gene. Serial dilu ons of DNA extracted fromE. coli host cells, along with DNA extracted from Active Pharmaceutical Ingredients of rSK, and IFN-alpha samples were subjected to an optimized real-time PCR assay based on SYBR Green chemistry
Results: The test enabled us to detect a small quantity of genomic DNA contamination as low as 0.0002 pg in recombinant protein-based drugs. For the first time, this study showed that DNA contamination in rSK and IFN-alpha preparation manufactured in Pasteur Institute of Iran is much lower than the safety limit suggested by the US FDA
Conclusion: Real-time PCR is a reliable test for rapid detection of host-cell DNA contamination, which is a major impurity of therapeutic recombinant proteins to keep manufacturers' minds on refining drugs, and provides consumers with safer biopharmaceuticals