Transplantation into NOD/SCID mice of ex vivo cultured subsets of AC133⁺and AC133⁻ cells isolated from the CD34⁻CD38⁻Lin⁻population. (A) Human-mouse DNA mixtures were used for PCR amplification of CART-1 human-specific gene sequence. A total of 200 ng of genomic DNA from BM cells of nontransplanted control NOD/SCID mice was mixed with equal volumes of serially diluted (0, 0.0625, 0.125, 0.25, and 0.5 ng) human genomic DNA. There were no detectable PCR products in the absence of human DNA, indicating the specificity of the PCR reaction to human sequence. However, increased amounts of human DNA allowed for the detection of a linearly increasing signal. PCR reactions of the human-mouse DNA mixture were compared to 200 ng DNA extracted from NOD/SCID mice transplanted with ex vivo–cultured cell populations as indicated. (B) Summary of the level of human cell engraftment detectable in NOD/SCID mice transplanted with AC133⁺ and AC133⁻ subfractions isolated from the CD34⁻CD38⁻Lin⁻populations in 8 independent experiments. Experiments 1 to 3 represent transplantation of NOD/SCID with uncultured de novo isolated subfractions, whereas experiments 4-8 represent results from mice transplanted with subfractions cultured for 3 days in serum-free media. Experiments 2, 3, and 8 represent transplantation of cells isolated from 3 pooled CB samples.