title: Human genome-specific real-time PCR method for sensitive detection and reproducible quantitation of human cells in mice.
abstract: Xenotransplantation of human cells into immunodeficiency mice has been frequently used to study stem cells in tissue repair and regeneration and cancer cell metastasis. However, a sensitive and reproducible method to quantify cell engraftment lacks. Here, we developed a Real-Time PCR-based method which facilitated consistent detection and quantification of small amounts of human cells distributed in mouse organs after infusion. The principle of the method was to directly detect a humans-specific sequence in the human-murine genomic DNA mixture. In a mouse myocardial infarction model, the Real-Time PCR-based method consistently determined the amounts of human mesenchymal stem cells (hMSCs) engrafted into the heart and other organs 7 days after infusion of as little as 2.5 × 10(5) cells, indicating a high sensitivity, and the amounts of hMSCs detected in mice highly correlated to the numbers of hMSCs transplanted. Importantly, different from previous PCR-based methods, our method produced highly consistent and reproducible results. The reliability of the method was further proven by parallel analyses of DiI-labeled hMSCs in tissue sections and in single cell suspensions of mice. Our data show that the present human genomic DNA-specific primers-based Real-Time PCR method is sensitive and highly reproducible in determining the amount of xenotransplanted human cells in murine tissues.
authors.affiliation: School of Life Sciences, Tsinghua University, Haidian, China.
authors.forename: Pengyue
authors.initials: P
authors.lastname: Song
authors.forename: Zhenhua
authors.initials: Z
authors.lastname: Xie
authors.forename: Ling
authors.initials: L
authors.lastname: Guo
authors.forename: Chengmei
authors.initials: C
authors.lastname: Wang
authors.forename: Weidong
authors.initials: W
authors.lastname: Xie
authors.forename: Yaojiong
authors.initials: Y
authors.lastname: Wu
mesh_terms: D000818:Animals; D002452:Cell Count / Q000379:methods*; D005260:Female; D015894:Genome, Human; D006801:Humans; D045164:Mesenchymal Stem Cell Transplantation; D059630:Mesenchymal Stem Cells / Q000166:cytology*; D051379:Mice; D008807:Mice, Inbred BALB C; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D014183:Transplantation, Heterologous