The Necessity for E.coli residual DNA testing
In fields such as cell and gene therapy, viral plasmids are amplified and fermented using E. coli as the host. The amplified plasmid needs to undergo E.coli residual DNA quality control before it can be used to infect 293 cells and package the virus. Therefore, the detection of E.coli residual DNA is critical.
Regulatory Requirements and Testing Methods
Current WHO and US FDA guidelines recommend residual DNA in finished products to be no greater than 10 ng/agent, and the US FDA also states that residual DNA in host cells of biologics should be no greater than 100 pg/agent. The General Principles of the European Pharmacopoeia stipulates that the limit of residual DNA in biological products should not be greater than 10 ng/dose, but the residual DNA limit for some particular vaccines is more stringent, e.g., the residual DNA in the inactivated vaccine against hepatitis A should not be higher than 100 pg/dose, and that in the vaccine against hepatitis B should not be higher than 10 pg/dose. The 2020 edition of the Chinese Pharmacopoeia, Part III, stipulates that the DNA residue in biological preparations produced on a cellular matrix should not exceed 100 pg/dose, and the DNA residue in vaccines produced on a bacterial or fungal matrix should not exceed 10 ng/dose.
In addition, for the methods of exogenous DNA residue determination, national pharmacopoeias also give guidance recommendations. The U.S. Pharmacopoeia 2017 edition of USP40-NF35 General Provision 1130 describes three methods for the determination of exogenous DNA residues, which are DNA probe hybridization, threshold method and real-time quantitative PCR method. The European Pharmacopoeia proposes real-time quantitative PCR and immunoenzymatic method, two sensitive analytical methods for quantifying host cell residual DNA. The Chinese Pharmacopoeia 2020 version of the three general rules 3407 also stipulates that the host cell DNA residue detection methods are DNA probe hybridization, fluorescence staining and quantitative PCR.
Among them, qPCR method has extremly high sensitivity, sequence specificity and accuracy, which can provide a reliable means of detection for the biopharmaceutical industry in process research and quality control of finished products, and it has now become the preferred detection method for each biologics manufacturers.
Information about Bluekit Products
Product Features
Rapid detection, strong specificity, reliable performance, the lowest detection limit can reach the “fg” level.
Product Parameters
- Detection range: 3.00×101 ~3.00×105fg/μL
- Limit of quantification：00×101fg/μL
- Detection limit：00 fg/μL
- Precision: CV% ≤ 15%.
Common Detection Problems and Precautions
1. This kit is for in vitro research use only, not for clinical diagnosis.
2. The kit must be used within the validity period.
3. All the components in the kit are recommended to be melted in a low-temperature environment and then used.
4. Only strictly comply with the instructions of the operation method, all the use of the kit supporting reagents to ensure the best detection effect.
5. Pay attention to the different sampling steps in a timely manner to replace the tip, to avoid cross-contamination, avoid opening the lid for a long time.
6. The final test results can be significantly impacted by the effectiveness of the reagents, the operator's operating methods and the test environment.
Product Consultation
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Email: info@hillgene.com
Post time: 2024-01-11 10:31:30
