Anti cd19 apc hib19

Twelve weeks after transplantation, mice were euthanized, and the bones were harvested from the mice (2× femur, 2× tibia, sternum, 2× pelvis, and spine) and crushed using a mortar and pestle. Mononuclear cells were enriched by using Ficoll gradient centrifugation (Ficoll-Paque Plus, GE Healthcare) for 25 min at 2,000 g at room temperature. After centrifugation, mononuclear cells were collected from the Ficoll layer and blocked to prevent nonspeci c antibody binding (TruStain FcX, BioLegend) and stained (30 min, 4 °C, dark) with monoclonal antibodies: anti-human CD45 V450 (HI30; BD Biosciences); anti-HLA-ABC APC-Cy7 (W6/32; BioLegend); anti-CD19 APC (HIB19; BD Biosciences); and anti-CD33 PE (WM53, BD Biosciences). Stained cells were then washed and resuspended in FACS buffer and analyzed on a BD FACS Sort II Aria. HLA-ABC+/CD45+ cells represent human engraftment. Normal multilineage engraftment was de ned by the presence of myeloid cells (CD33+) and B-cells (CD19+) within engrafted human CD45+HLA-ABC+ cells. GFP expression within engrafted human CD45+HLA-ABC+ cells was also analyzed by ow cytometry and represents the percentage of targeted cell engraftment.

Twelve weeks after transplantation, mice were euthanized, and the bones were harvested from the mice (2× femur, 2× tibia, sternum, 2× pelvis, and spine) and crushed using a mortar and pestle. Mononuclear cells were enriched by using Ficoll gradient centrifugation (Ficoll-Paque Plus, GE Healthcare) for 25 min at 2,000 g at room temperature. After centrifugation, mononuclear cells were collected from the Ficoll layer and blocked to prevent nonspeci c antibody binding (TruStain FcX, BioLegend) and stained (30 min, 4 °C, dark) with monoclonal antibodies: anti-human CD45 V450 (HI30; BD Biosciences); anti-HLA-ABC APC-Cy7 (W6/32; BioLegend); anti-CD19 APC (HIB19; BD Biosciences); and anti-CD33 PE (WM53, BD Biosciences). Stained cells were then washed and resuspended in FACS buffer and analyzed on a BD FACS Sort II Aria. HLA-ABC+/CD45+ cells represent human engraftment. Normal multilineage engraftment was de ned by the presence of myeloid cells (CD33+) and B-cells (CD19+) within engrafted human CD45+HLA-ABC+ cells. GFP expression within engrafted human CD45+HLA-ABC+ cells was also analyzed by ow cytometry and represents the percentage of targeted cell engraftment.