Chinese Hamster Ovary Cell Genome DNA Residue Reference Material for Testing

LIANGWen; CAO Mei-xia; LIU Gang

doi:10.3969/j.issn.1000-1158.2025.05.19

PDF(1686 KB)

Acta Metrologica Sinica ›› 2025, Vol. 46 ›› Issue (5) : 754-761. DOI: 10.3969/j.issn.1000-1158.2025.05.19

Chinese Hamster Ovary Cell Genome DNA Residue Reference Material for Testing

Author information +

History +

Abstract

To solve the traceability problem of the exogenous DNA residue determination method （qPCR method） in Chinese Pharmacopoeia （2020 edition）， a large-scale cultivation of Chinese hamster ovary （CHO） cells was conducted. Various techniques such as column chromatography， ethanol precipitation， and ultrafiltration were employed to extract high-purity CHO genomic DNA. Digital PCR， which has been extensively studied and optimized， was chosen as the quantitative method for genomic DNA. The results showed that the digital PCR-based method effectively overcame matrix interference issues and demonstrated better consistency than traditional UV detection. The uniformity and stability of the standard substance were verified， and the copy number concentration was determined using 5 different digital PCR platforms from 10 units， resulting in a final value of （1.06±0.12）×10⁴copies/μL. This standard substance can facilitate the development of DNA residual testing kits and promote mutual recognition and standardization of DNA residual testing results among biopharmaceutical companies， ensuring traceability of the testing results.

Key words

biometrology / CHO cell / genome DNA / residual detection / digital PCR / reference materials

Cite this article

EndNote

Ris (Procite)

Bibtex

Download Citations

LIANGWen , CAO Mei-xia , LIU Gang. Chinese Hamster Ovary Cell Genome DNA Residue Reference Material for Testing[J]. Acta Metrologica Sinica. 2025, 46(5): 754-761 https://doi.org/10.3969/j.issn.1000-1158.2025.05.19

References

List( Publishing order | Descend order by publishing year | Descend order by cited within ) Chart analysis

1	侯慧丽，王艺诺，汤晓闯，等.重组人凝血因子Ⅷ产品中CHO细胞DNA残留量的检测［J］.中国生物制品学杂志， 2018，31（6）：667-670. HOU H L， WANG Y N， TANG X C， et al.Detection of Residual CHO Cell DNA in Recombinant Human Coagulation Factor VIII Products［J］.ActaMetrologica Sinica， 2018， 31（6）： 667-670. Cited in this article [1]

刘莹，刘玉林，俞露，等.长效重组人生长激素纯化液中CHO-K1细胞残留DNA实时荧光定量PCR检测方法的建立及验证［J］.中国生物制品学杂志， 2019，32（06）：688-693.

LIU

， LIU

Y L

， YU

， et al. Development and Validation of a Real-Time Fluorescence Quantitative PCR Method for Detecting Residual CHO-K1 Cell DNA in the Purification Solution of Long-Acting Recombinant Human Growth Hormone［J］.Chinese Journal of Biologicals，2019，32（6）：688-693.

3	祁自柏，米山徹夫，宫村逵男. 乙肝基因工程疫苗中宿主细胞DNA的检测［J］. 中华微生物学和免疫学杂志， 1989，9（4）：255-257，271. QI Z L， Tetetsuo Y， Miyamura K. Detection of Host Cell DNA in Hepatitis B Genetic Engineering Vaccine［J］.Chinese Journal of Microbiology and Immunology，1989，9（4）：255-257，271.

4	GENG S L， ZHAO X J， ZHANG X， et al. Recombinant therapeutic proteins degradation and overcoming strategies in CHO cells［J］. Applied Microbiology and Biotechnology， 2024， 108（1）：182.

5	宋雨蒙，翟睿，雷喜梅，等.肿瘤细胞外泌体的生物学功能及其检测方法研究进展［J］.计量学报，2023，44（06）：992-999. SONG Y M，ZHAIR， LEI X M， et al.Research Advances in the Biological Functions and Detection Methods of Tumor Cell-Derived Exosomes［J］.ActaMetrologica Sinica，2023，44（6）：992-999. Cited in this article [1]

6	ZHANG J H， SHAN L L， LIANG F， et al. Strategies and considerations for improving recombinant antibody production and quality in Chinese hamster ovary cells［J］. Frontiers in Bioengineering and Biotechnology， 2022， 10：856049. Cited in this article [1]

7	TIHANYI B， NYITRAY L. Recent advances in CHO cell line development for recombinant protein production［J］. Drug Discovery Today： Technologies， 2020， 38： 25-34. Cited in this article [1]

8	LI F， LIU B， ONG Y， et al.Enhanced Downstream Processing for a Cell-Based Avian Influenza （H5N1） Vaccine［J］. Vaccines， 2024， 12（2）：138. Cited in this article [1]

9	YANG D， HUANG L， WANG J， et al.Tumorigenesis mechanism and application strategy of the MDCK cell line： A systematic review［J］. Biologicals， 2023， 83：101699. Cited in this article [1]

KNEZEVIC

， STACEY

， PETRICCIANI

， et al. Evaluation of cell substrates for the production of biologicals： Revision of WHO recommendations： Report of the WHO Study Group on Cell Substrates for the Production of Biologicals， 22–23 April 2009， Bethesda， USA［J］. Biologicals， 2010， 38（1）：162-169.

Cited in this article [1]

EMA. Position Statement on the Use of Tumourigenic Cells of Human Origin for the Production of Biological and Biotechnological Medicinal Products， The Euro-pean Agency for the Evaluation of Medicinal Products： Evaluation of MedicinalProducts for Human Use［S］Amsterdam：EMA，2001.

Cited in this article [1]

12	FDA. Guidance for Industry： Characterization and Qualification of Cell Substrates and Other Biological Materials Used in the Production of Viral Vaccines for Infectious Disease Indications， US Food and Drug Administration ［S］.USA：FDA，2010. Cited in this article [1]

13	LI D， ZHANG Q， LIU G， et al. Detection of residual E. coli host cell DNA by 23S ribosomal RNA gene-targeted quantitative polymerase chain reactions［J］. Journal of Pharmaceutical and Biomedical Analysis， 2021， 198： 114000. Cited in this article [1]

郑子繁，王颢潜，高佳奇，等.耐除草剂转基因大豆SHZD32-1数字PCR绝对定量方法的建立［J］.计量学报，2023，44（3）：415-421.

ZHENG

Z F

， WANG

H Q

， GAO

J Q

，et al. Development of an Absolute Quantification Method Using Digital PCR for Herbicide-Tolerant Genetically Modified Soybean SHZD32-1［J］. ActaMetrologica Sinica，2023，44（3）：415-421.

Cited in this article [1]

15	The United States Pharmacopieial Convention. The United States Pharmacopeia （43th Edition）： General Chapters ［S］.USA：United States Pharmacopeial Convention，2020 Cited in this article [2]

16	国家药典委员会.中华人民共和国药典2020 年版（四部）［M］. 北京：中国医药科技出版社， 2020 Cited in this article [1]

17	国家市场监督管理总局. 生物技术核酸靶序列定量方法的性能评价要求 qPCR法和dPCR法：GB/T 42077-2022 ［S］. 北京：中国标准出版社出版，2022. Cited in this article [1]