Figure 8.
Figure 8. Assessment of engraftment of BCR-ABL-positive cells transplanted into NOD/SCID recipient mice. Twelve days after tetracycline removal from the drinking water, cells were isolated from the bone marrow of a single-transgenic (sTG) or a double-transgenic (dTG) BCR-ABL-positive donor mouse and were transplanted into sublethally (300 cGy) irradiated NOD/SCID mice (recipient 9 for sTG; recipients 11A, 11B, 13 for dTG). At this time point, the dTG donor mouse showed neutrophilia, anemia, splenomegaly, and granulocytic hyperplasia of the bone marrow and spleen. In separate experiments, bone marrow-derived FACS-sorted Lin-c-kit+Sca-1+ HSCs from pooled, diseased dTG donor mice were transplanted into a 300 cGy-irradiated NOD/SCID mouse (recipient 8), and lymphoblastic cells from an affected lymph node of a dTG mouse were transplanted into a nonirradiated NOD/SCID mouse (recipient 14). As indicated, 70 or 105 days after transplantation, genomic DNA was isolated from the bone marrow of the recipients. BCR-ABL genomic DNA was amplified by real-time PCR and expressed as the percentage of the amount of amplified murine CCAAT enhancer binding protein alpha (mCEBPA) genomic DNA to quantify the engraftment of BCR-ABL-positive donor cells. Genomic DNA from the bone marrow of a wild-type NOD-SCID mouse that did not undergo transplantation and from the tails of 2 wild-type (wt) and 2 SCLtTA/BCR-ABL (dTG) mice served as negative and positive controls. None of the recipient mice have shown any signs of disease within the 5-month observation period.

Assessment of engraftment of BCR-ABL-positive cells transplanted into NOD/SCID recipient mice. Twelve days after tetracycline removal from the drinking water, cells were isolated from the bone marrow of a single-transgenic (sTG) or a double-transgenic (dTG) BCR-ABL-positive donor mouse and were transplanted into sublethally (300 cGy) irradiated NOD/SCID mice (recipient 9 for sTG; recipients 11A, 11B, 13 for dTG). At this time point, the dTG donor mouse showed neutrophilia, anemia, splenomegaly, and granulocytic hyperplasia of the bone marrow and spleen. In separate experiments, bone marrow-derived FACS-sorted Lin-c-kit+Sca-1+ HSCs from pooled, diseased dTG donor mice were transplanted into a 300 cGy-irradiated NOD/SCID mouse (recipient 8), and lymphoblastic cells from an affected lymph node of a dTG mouse were transplanted into a nonirradiated NOD/SCID mouse (recipient 14). As indicated, 70 or 105 days after transplantation, genomic DNA was isolated from the bone marrow of the recipients. BCR-ABL genomic DNA was amplified by real-time PCR and expressed as the percentage of the amount of amplified murine CCAAT enhancer binding protein alpha (mCEBPA) genomic DNA to quantify the engraftment of BCR-ABL-positive donor cells. Genomic DNA from the bone marrow of a wild-type NOD-SCID mouse that did not undergo transplantation and from the tails of 2 wild-type (wt) and 2 SCLtTA/BCR-ABL (dTG) mice served as negative and positive controls. None of the recipient mice have shown any signs of disease within the 5-month observation period.

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