Donor stromal cells from human blood engraft in NOD/SCID mice by Silvia-Renate Goan, Ilse Junghahn, Manuela Wissler, Michael Becker, Jutta Aumann, Ursula Just, Georg Martiny-Baron, Iduna Fichtner, and Reinhard Henschler Blood Volume 96(12):3971-3978 December 1, 2000 ©2000 by American Society of Hematology

Human progenitor cells from peripheral and cord blood initiate a lymphomyelopoiesis in NOD/SCID mice.Single-cell suspensions were prepared from bone marrow of NOD/SCID mice transplanted with CD34-enriched PBCs or light-density CBCs, incubated with fluoresce... Human progenitor cells from peripheral and cord blood initiate a lymphomyelopoiesis in NOD/SCID mice.Single-cell suspensions were prepared from bone marrow of NOD/SCID mice transplanted with CD34-enriched PBCs or light-density CBCs, incubated with fluorescence-tagged monoclonal antibodies directed to human cell surface antigens, and analyzed by flow cytometry. Dot-plot analyses of dual marker-labeled cell preparations were performed. The percentages given in the panels represent the percentage of cells in the total ungated population (10 000 events). Representative analyses of the experiments in Table 1 are shown. Silvia-Renate Goan et al. Blood 2000;96:3971-3978 ©2000 by American Society of Hematology

Immunostaining of stromal marker-positive cells Immunostaining of stromal marker-positive cells.Immunostaining is shown for human-specific cell surface markers in stromal layers of chimeric bone marrow cultures (A-G) or cell suspensions (H,I,K) from NOD/SCID mouse bone marrow harvested 5 to 11 weeks afte... Immunostaining of stromal marker-positive cells.Immunostaining is shown for human-specific cell surface markers in stromal layers of chimeric bone marrow cultures (A-G) or cell suspensions (H,I,K) from NOD/SCID mouse bone marrow harvested 5 to 11 weeks after transplantation with human blood cells. Mice were transplanted with human CD34-enriched PBCs (A,B,D,H,I,J) or light-density CBCs (C,D,G). The antibodies used were directed to the following human cell surface antigens: (A) HLA class-I, (B) 5B5 (fibroblast), (C) vWF, (D,E) KDR/flk-1, (F) HLA class-I, (G) CD34, (H) HLA class-I, (I) AS02 (fibroblast), (K) HLA class-I. Positive staining appears as red or brown. Controls from bone marrow from nontransplanted mice were processed in parallel for all markers (F,K). (Original magnification, 40-fold [A], 400-fold [B-G], 1000-fold [H-K].)‏ Silvia-Renate Goan et al. Blood 2000;96:3971-3978 ©2000 by American Society of Hematology

RT-PCR amplification of RNA from mice RT-PCR amplification of RNA from mice.RNA was isolated from the bone marrow or spleen cell suspensions of transplanted or nontransplanted mice as indicated. RT-PCR amplification of RNA from mice.RNA was isolated from the bone marrow or spleen cell suspensions of transplanted or nontransplanted mice as indicated. The cDNA was prepared and analyzed for the presence of transcripts for human KDR or human proline hydroxylase as described in “Patients, materials, and methods.” The PCR products were separated on agarose gels and visualized with ethidium bromide. Negative control, mock PCR using water instead of cDNA; positive control, cDNA from human umbilical vein endothelial cells (HUVECs) for KDR, or KMST-6 fibroblasts of human Dexter cultures for proline hydroxylase. CD45+CD14+: analysis of sorted cells from Dexter cultures as described in “Patients, materials, and methods.” A total of 3 of 9 transplanted mice were positive for human KDR cDNA and 6 of 8 mice for proline hydroxylase. BMC indicates analysis from a bone marrow culture established from the bone marrow of a mouse transplanted with CD34+ PBCs. Silvia-Renate Goan et al. Blood 2000;96:3971-3978 ©2000 by American Society of Hematology