Cd34 bv785 clone 561

The stromal vascular fraction was labeled with 2 μl of controls or fluorophore-conjugated antibodies in Eppendorf tubes at room temperature for 20 minutes. After that, the cells were fixed, and the erythrocytes were lysed with 1 ml of BD FACS Lysing Solution for 30 minutes. Then, the samples were centrifuged 10 minutes at 3500 x g, and the pellets were resuspended in 500 μl of PBS. Subsequently, the samples were stored at 4°C until the next day. Flow cytometry was performed using a FACS ARIA III equipment, and data were acquired on a logarithmic scale. The internal standard was used to calculate the number of cells per mg of tissue.
The fluorescent-conjugated antibodies used to identify mast cells were: anti-CD45 PE-CF594 (clone HI30, BD), anti-CD117 APC (clone YB5.B8, BD), anti-FcϵRI PE-Cy7 (clone AER-37, BioLegend), and CD203c BV421 (clone NP4D6, BioLegend). Additionally, the antibodies CD34 BV785 (clone 561, BioLegend) and integrin β7 FITC (clone FIB504, BioLegend) were used in a subcohort of 15 patients (6 non-T2D, 6 pre-T2D, and 3 T2D). Compensation beads and isotype controls were purchased from BD Biosciences. MC were identified as CD45⁺ CD117⁺ CD203c⁺ FcϵRIα⁺ (Figure 1A).

Cynomolgus macaques^{37 (link)} were treated with intravenous injection of CD117-ADC (0.1 or 0.3 mg/kg × 1 day) or busulfan (6 mg/kg × 4 days), and bone marrow CD34 + CD90 + CD45RA- cells were quantified by using antibodies against CD34 BV785 (clone 561, BioLegend, 1:300), CD90 APC (clone 5E10, BioLegend, 1:300), and CD45RA VioBlue (clone T6D11, Miltenyi Biotec, 1:50) in flow cytometry 7 days after drug administration (n = 3 per group) and compared to historical phosphate-buffered saline (PBS)-treated naïve animals. Bone marrow cellularity was assessed by complete blood count analysis to determine white blood cells per ml of marrow. Flow cytometry was used to determine the frequency of CD34 + CD90 + CD45RA- cells of total CD45+ cells. The absolute number of CD34 + CD90 + CD45RA- was determined by multiplying the frequency of CD34 + CD90 + CD45RA- cells (out of total CD45 cells) with the bone marrow cellularity in cells per ml.