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Duplex Polymerase Chain Reaction Quantification of Human Cells in a Murine Background

^a Laboratory of Molecular Medicine, Interdisciplinary Centre for Clinical Research, University of Leipzig Faculty of Medicine, Germany;
^b In Vitro Differentiation Group, IPK, Gatersleben, Germany;
^c Institute for Immunology, University of Leipzig Faculty of Medicine, Germany;
^d Vita 34 AG, Leipzig, Germany

Key Words. Species-specific polymerase chain reaction • Human • Mouse • Chimera • Stem cell • Metastasis • Quantification

Correspondence: Michael Cross, Ph.D., Division of Hematology/Oncology, University of Leipzig, Inselstrasse 22, 04103 Leipzig, Germany. Telephone: 49-0-341-97-15942; Fax: 49-0-341-97-15979; e-mail: crossm{at}medizin.uni-leipzig.de

Studies of the regenerative potential of human stem cells commonly involve their transplantation into immune-deficient mice or in vitro coculture with mouse cells. The optimal use of such models requires the detection and quantification of relatively low numbers of human cells in a murine background. We report here a duplex polymerase chain reaction (PCR) approach involving the coamplification of human-and mouse-specific repetitive sequences. The determination of product ratios compensates against variations in sample quality and enables quantitation from >50% down to 0.01% human-in-mouse from a single reaction. Product ratios are determined by standard electrophoresis of end-stage PCR reactions followed by image analysis techniques using freely available software, with no requirement for real-time PCR. The approach has been used to analyze tissue from mice transplanted with human cells and cocultures between differentiating mouse embryonal stem cells and human umbilical cord blood cells.