ABSTRACT: Objective To establish a quantitative PCR method for the analysis of human-derived SRY gene DNA in mouse tissues, and to study the tissue distribution of human umbilical cord mesenchymal stem cells in immunodeficient NOG mice after a single intravenous injection. Method A quantitative PCR method for the analysis of human SRY gene DNA in mouse tissues was established, and the methodological validation of standard curve and linear range, accuracy and precision, and stability was performed. Thirty-six NOG mice, one half of each sex, were given HUCMSCs 3.5?07 cells/kg by single intravenous injection, and six mice were anesthetized at 6 h, 12 h, 24 h, 72 h, 1 w and 2 w, respectively, and dissected after blood collection (EDTA anticoagulation). DNA was extracted from lung, kidney, heart, liver, brain, spinal cord, stomach, small intestine, fat, skin, spleen and testis (uterus and ovary), and the distribution of HUCMSCs in each tissue was determined by the validated method of quantitative PCR analysis of human-derived SRY gene in mouse tissues. In addition, 18 NOG mice, half males and half females, were divided into control group (6 mice) and treating group (12 mice), which were injected intravenously with 0.9% sodium chloride injection and HUCMSCs 3.5?07 cells/kg. The acute toxic reactions of mice were observed during the administration period, and four animals were dissected at 72 h, 2 w and 4 w after the administration to observe the gross organs, and the mitochondrial protein expression was detected by immunohistochemistry in paraffin sections of lung tissues to analyze the colonization of HUCMSCs in lung tissues. Results The established quantitative PCR method for human-derived SRY gene DNA in mouse tissues met the validation criteria for each index. After a single intravenous injection in NOG mice, HUCMSCs were mainly distributed in the lungs and bloodwithin 1w after administration, with higher concentrations in lung tissues than in blood, and the concentration of HUCMSCs in lung tissue and blood maintained relatively stable levels within 6h~24h and 6h~72h, respectively, and then decreased over time. The distribution of HUCMSCs was not measured in other tissues at all sampling points . The colonization results showed that HUCMSCs were detected in lungs 72h after intravenous injection, but not at 2w and 4w. No obvious acute toxicity was observed in NOG mice after single intravenous administration of HUCMSCs. Conclusion The established method for analyzing the distribution of HUCMSCs in mouse tissue is reliable and feasible. HUCMSCs were mainly distributed in lung and blood of NOG mice within 1w after single intravenous injection, and mainly colonized in lung tissue at 72h. The single intravenous administration of HUCMSCs has a good safety profile.