Application of Rapid and Sensitive Real Time PCR Technique in Detection of DNA Impurities in Recombinant Interferon

Abstract

Background & Objective: Interferon belongs to a family of cytokines, which has the most important role in the innate immune response to virus infections. While producing recombinant interferon in biological host, some pieces of host nucleic acids remain in product. Because of limitations in previous techniques for detection of these impurities, the objective of this study is to use rapid and sensitive Real time PCR method for detecting the impurities. Materials & Methods: First, with DNA extraction from bacterial host cell and preparation of its serial dilutions, SYBR Green-based Real time PCR reaction was held and standard curve was plotted. After DNA extraction from interferon and performing PCR, total DNA amount was determined using standard curve. Results: Studies performed on some interferon samples, revealed that the amount of DNA impurities was about 0.02 pg. per product dose. In addition, the designed primers in the above reaction had no interaction with each other and other interfering agents. Conclusion: For the first time in Iran, this study was set up and it revealed that Real time PCR can be used as a functional and accurate technique in manufacture centers for detection of residual host cell DNA in interferon and other recombinant pharmaceutical products.

Keywords

• dna impurities
• recombinant pharmaceutical products
• interferon
• real time pcr
• sybr green i