Etv6^+/RUNX1 mice with “second hits” develop a pre-B ALL recapitulating features of the human disease. (A) Peripheral blood (PB; original magnification ×2000), spleen, bone marrow, and liver (original magnification ×400 in top panels and original magnification ×1000 in bottom panels) from a representative mouse with BCP-ALL. The presence of lymphoblasts in the PB is obvious, as is the infiltration of the spleen, bone marrow and liver, with effacement of the normal cellular architecture and replacement by nucleolated blasts. (B) FACS plots from the bone marrow of a representative mouse demonstrate only background Gr-1/Mac1 myeloid cells, with the majority of cells having a B220⁺/CD19⁻/sIg⁻ phenotype in keeping with BCP-ALL. (C) D-J PCR rearrangement studies (involving DQ52 and DFL/DSP genes) were performed on spleen or bone marrow gDNA of 3 mice with phenotypic B-ALL. Negative control was gDNA from C57BL/6J mouse kidney (known to be unrearranged), and positive control was gDNA from a C57BL/6J primary pro-B cell bone marrow culture (known to contain a lot of DJ-rearranged alleles). Sample 1 shows 2 different rearrangement events (DQ52-J2 and DFL/DSP-J2), in keeping with a clonal B-cell population with both alleles D-J recombined. Sample 2 shows both a germ line band and an identical DFL/DSP-J2 band, in keeping with a clonal B-cell population with rearrangement of one allele and germ line configuration of the other. Sample 3 shows an absence of recombination.