Anti human cd45 percp

Prior to the analyses, erythrocytes were lysed with Red Blood Lysis Buffer (154 mM NH₄Cl, 14 mM NaHCO₃ and 0.1 mM EDTA). For flow cytometry analyses the following reagents were used: FcRBlocking Solution (BD Biosciences 553142); anti Human CD45 PerCp (BD Bioscience BMS45-9459-42); anti Human CD3 PE (BD Biosciences 555340); anti Human CD19 FITC (BD Biosciences 555412); anti Human CD33 PE (BD Biosciences 555450); anti Human CD11b FITC (BD Biosciences 562793) and, as an isotipic control, Simultest (BD Biosciences 342409). Flow-cytometric analysis was performed using the FACScan Beckton Dickinson and FlowJoX software. Blood cell counts were performed with the VetScanHM5 (ABAXIS). After 12 weeks, four mice of each cohort were sacrificed by cervical dislocation. Bone marrow from femur and tibia of these animals was sampled as described by Soleimani and Nadri [72 (link)] and collected in DMEM 10% FBS, 10 U/ml Heparin for flow-cytometry analysis and secondary transplant.
To monitor the secondary engrafment, flow-cytometric analyses of human CD45⁺ cells in the peripheral blood of the secondary recipients were performed 9 and 14 weeks after the transplant. On week 16, the mice were sacrificed and their bone marrow was collected and analysed.