Intrinsically reduced function of Vav-Cre Rb KO HSPCs outside the BM and upon transplantation. (A) Frequency/function of HPCs and HSCs analyzed by the CAFC assay. Shown is the experimental setup of the assay. (B) Frequency/function of distinct CAFC populations in Vav-Cre Rb KO animals in BM relative to control animals analyzed in parallel (n = 6 for control and Vav-Cre Rb KO). The dotted line (value of 1) indicates the level of no relative difference in CAFCs between Vav-Cre Rb KO and control animals in this figure. Starting from CAFC day 21 on, Vav-Cre Rb KO primitive hematopoietic cells are significantly reduced in their frequency/function. (C) Analysis of the progenitor cell function of Vav-Cre Rb KO cells upon transplantation of BM cells into sublethally irradiated NOD/SCID recipients. Shown is the experimental setup. (D) Determination of the contribution of control and Vav-Cre Rb KO cells to PB 3 weeks and 6 weeks after transplantation (chimerism in PB) by flow cytometric analyses (n = 4 recipients for control and n = 4 recipients for Vav-Cre Rb KO). (E) Analysis of stem- cell function by competitive repopulation assays into C57BL/6 CD45.1 animals. Shown is the experimental setup. (F) Determination of the contribution of control and Vav-Cre Rb KO cells to PB up to 27 weeks after transplantation and up to 6 weeks in secondary recipients (chimerism in PB) by flow cytometric analyses (n = 3 recipients for control and n = 3 recipients for Vav-Cre Rb KO). Shown are mean values plus or minus 1 SEM; *P < .05.