Pe anti human cd34 antibody

The standard chemicals, reagents, and the specific antibodies were purchased from Sigma Chemical Co. (St. Louis, MO, USA), Santa Cruz Biotechonolgy (European Support Office, Heidelberg, Germany), Merk & Co (Kenilworth, NJ, USA), Lonza (Basel, Switzerland), R&D Systems (Minneapolis, MN, USA), Qiagen (Hilden, Germany), and from Thermo Fisher Scientific (Waltham, MA, USA). All others reagents were of analytical grade.
For flow cytometry used the following antibodies: Annexin V(FL)-FITC (Santa Cruz, sc-4252 FITC, anti-human, source: E. coli, c = 0.05 µg/µL); PE anti-human CD34 antibody (Biolegend, cod 343606, host species: mouse, c = 0.005 µg/µL); CD133/2(293C3)-APC antibody (Millteny, Order no. 130-090-854, anti-human, clone: 293C3, c = 10 µL/10⁶ cells); human VEGFR2/KDR/Flk-1 Fluorescein-conjugated antibody (R&DSystem, FAB357F, source: monoclonal mouse IgG1 clone # 89106, c = 10 µL/10⁶ cells); Goat F(ab’)2 Anti-Human IgG H&L- (Phycoerythrin) pre-adsorbed antibody (abcam-ab7006, host species: goat, dilution: 1/100); goat anti-human IgG (H+L) cross-adsorbed secondary antibody, FITC (Invitrogen-31531, host species: goat, dilution: 1/100).

CAFs were processed to cell suspension and counted. The antibodies were added as manufacturer’s guidance (FITC anti-human CD105 Antibody, Biolegend, USA, Cat: 323203. FITC anti-human CD90 Antibody, Biolegend, USA, Cat: 328107. PE anti-human CD146 Antibody, Biolegend, USA, Cat: 361005. PE anti-human CD34 Antibody, Biolegend, USA, Cat: 378603. PE anti-human HLA-DR Antibody, Biolegend, USA, Cat: 327007. PE anti-human CD73 Antibody, Biolegend, USA, Cat: 344003). After incubated for 30 min at 37℃ in dark conditions, the samples were analyzed or sorted. Then the cells were collected and cultured in DMEM/F12 media supplemented with 10% FBS and 1% penicillin/streptomycin.