Evidence map›Paper›PMID 37805662›Full record

ArticleMolecular biology reports2023

An efficient qPCR assay for the quantification of human cells in preclinical animal models by targeting human specific DNA in the intron of BRCA1.

Ke Liu, Lang Hu, Siyu Wang, Xinzhu Chen, Yuting Liu, Shuli Zhao, Hui Wang, Li Li, Hui Li

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Article in Molecular biology reports, 2023. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Not yet cited in PubMed.

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5 · Who and what money

Authors and funding

9 authors.

Ke Liu *Department of Gastroenterology, The Affiliated Hospital of Xuzhou Medical University, Xuzhou, 221000, Jiangsu, China.
Lang Hu *Jiangsu Key Laboratory of Immunity and Metabolism, Department of Pathogenic Biology and Immunology, Xuzhou Medical University, 209 Tongshan Road, Xuzhou, 221004, Jiangsu, China.
Siyu Wang *Jiangsu Key Laboratory of Immunity and Metabolism, Department of Pathogenic Biology and Immunology, Xuzhou Medical University, 209 Tongshan Road, Xuzhou, 221004, Jiangsu, China.
Xinzhu ChenJiangsu Key Laboratory of Immunity and Metabolism, Department of Pathogenic Biology and Immunology, Xuzhou Medical University, 209 Tongshan Road, Xuzhou, 221004, Jiangsu, China.
Yuting LiuJiangsu Key Laboratory of Immunity and Metabolism, Department of Pathogenic Biology and Immunology, Xuzhou Medical University, 209 Tongshan Road, Xuzhou, 221004, Jiangsu, China.
Shuli ZhaoGeneral Clinical Research Center, Nanjing First Hospital, Nanjing Medical University, Nanjing, China.
Hui WangJiangsu Key Laboratory of Immunity and Metabolism, Department of Pathogenic Biology and Immunology, Xuzhou Medical University, 209 Tongshan Road, Xuzhou, 221004, Jiangsu, China.
Li LiDepartment of Gastroenterology, The Affiliated Hospital of Xuzhou Medical University, Xuzhou, 221000, Jiangsu, China. lily9711214@126.com.
Hui LiJiangsu Key Laboratory of Immunity and Metabolism, Department of Pathogenic Biology and Immunology, Xuzhou Medical University, 209 Tongshan Road, Xuzhou, 221004, Jiangsu, China. lihuishenghua@xzhmu.edu.cn.

Funding

National Natural Science Foundation of China 82171791the Postgraduate Research & Practice Innovation Program of Jiangsu Province KYCX21_2638the Youth Innovation Team Grant and the Starting Grant by Xuzhou Medical University D2018009Xuzhou Science and Technology Program KC20089
6 · The paper itself

Abstract

backgroundPrecise quantification of grafted human cells in preclinical animal models such as non-human primates, rodents and rabbits is needed for the evaluations of the safety and efficacy of cell therapy. Quantitative PCR (qPCR) as a swift, sensitive and powerful assay is suitable for human cell quantification. However, it is a formidable challenge due to that the genome of non-human primates share more than 95% of similarity as human.

methodsIn the present study, we developed a probe-based quantitative PCR (qPCR) assay for the quantification of human cells in preclinical animal models via targeting human specific DNA in the intron of BRCA1 (termed BRCA1-qPCR). The 5' and 3' end of BRCA1-qPCR probe was conjugated with FAM and non-fluorescent quencher-minor groove binder (NFQ-MGB), respectively. 1 µg of genomic DNA from human and preclinical animal models including rhesus monkeys, cynomolgus monkeys, New Zealand white rabbits, SD rats, C57BL/6 and BALB/c mice were used for determining the specificity and sensitivity of the BRCA1-qPCR assay. A calibration curve was generated by BRCA1-qPCR analysis of linearized plasmid containing targeted human specific DNA in BRCA1. The BRCA1-qPCR assay was validated by analysis of 0.003%, 0.03% and 0.3% of human leukocytes mixed within murine leukocytes.

resultsThe BRCA1-qPCR assay detected human DNA rather than DNA from tested species. The amplification efficiency of the BRCA1-qPCR assay was 95.4% and the linearity of the calibration curve was R

conclusionThe BRCA1-qPCR assay is efficient for the quantification of human cells in preclinical animal models.

Indexed as

DNAPrimatesAnimalsBRCA1 ProteinHumansIntronsMiceMice, Inbred C57BLModels, AnimalPolymerase Chain ReactionRabbitsRatsRats, Sprague-DawleyBRCA1 ProteinBRCA1 protein, humanDNABRCA1Cell transfer therapyPreclinical animal modelsQuantitative PCR

Identifiers

PMID37805662

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