Simple and Efficient Isolation of Hematopoietic Stem Cells from H2K-zFP Transgenic Mice

¹ Department of Life Science, Swiss Federal Institute of Technology, CH-1015, Lausanne, Switzerland
² Department of Pathology and Developmental Biology, Stanford University School of Medicine, Stanford, California
³ Department of Life Science, Swiss Federal Institute of Technology, Lausanne, Switzerland; The Burnham Institute, 10901 North Torrey Pines Rd, La Jolla, California

^* To whom correspondence should be addressed. E-mail: terskikh{at}burnham.org.

	Abstract

We have generated a transgenic mouse line, which allows for simple and highly efficient enrichment for mouse hematopoietic stem cells (HSCs). The transgene expresses a GFP variant (zFP) under the control of H2K^b promoter/enhancer element. Despite the broad z FP expression, transgenic HSCs express exceptionally high levels of zFP, al-lowing prospective isolation of a population highly enriched in HSCs by sorting the 0.2% of the brightest green cells from the enriched bone marrow of H2K-zFP mice. Up to 90% of zFP^bright cells are also c-kit^high, Sca-1^high, Lin^neg, Flk-2^neg, which is a bona fide pheno-type for long-term HSCs. Double sorted zFP^bright HSCs were capable of long-term multi-lineage reconstitution at a limiting dilution dose of about 12 cells, which is comparable to that of highly purified HSCs obtained by conventional multicolor flow cytometry. Thus, the H2K-zFP transgenic mice provide a straightforward and easy setup for the simple and highly efficient enrichment for genetically-labeled HSCs without using fluorescence-conjugated monoclonal antibodies. This approach will greatly facilitate gene transfer, in-cluding siRNA for gene knock-down, into HSCs and, consequently, into all other hema-topoietic lineages.