Anti cd106

Isolation and identification of rat TSCs were performed as previously described [16 (link)]. Briefly, 0.3% pentobarbital sodium (Sigma, 30 mg/kg) was used for intraperitoneal anesthesia in rats. In sterile conditions, the tendon tissues were then removed, carefully dissected, cut into pieces, and digested in 3 mg/mL of type I collagenase (Sigma-Aldrich, St. Louis, MO, USA). After a 70-μm cell filter filtration, the suspension turned into a single cell suspension, which was then cultured in Dulbecco’s modified Eagle’s medium (Gibco, Invitrogen, NY, Invitrogen Corporation, Grand Island, USA) containing 10% fetal bovine serum (Biological Industries, Kibbutz Beit-Haemek, Israel) and 1% penicillin-streptomycin antibiotic mixture (Beyotime, Shanghai, China). Cells were subcultured at 80% confluence. Cells at passages three were incubated with fluorescein isothiocyanate-conjugated antibodies (anti-CD90, anti-CD105, anti-CD44, anti-CD11b, and anti-CD106) (Biolegend San Diego, CA, USA) through flow cytometry. The multilineage differentiation potential of TSCs was determined by inducing the differentiation of cells in passage 3 into osteocytes, adipocytes, and chondrocytes (all the differentiation media were from Cyagen).

AT-MSCs and CB-MSCs (2 × 10⁴ cells) were seeded into 6-well plates on day 0. The total number of expanded cells were counted at day 1, 3 and 5. Cell doubling time (DT) was calculated by the following formula: DT = T × ln2/ln × (Xe/Xb), where T is the incubation time in any units; Xb is the cell number at the beginning of the incubation time; Xe is the cell number at the end of the incubation time.
Cultured cells were stained with FITC-conjugated anti-CD29 or anti-c-kit, PE-conjugated anti-CD44, anti-CD45, anti-CD106 or anti-Sca-1, APC-conjugated anti-CD105, and APC-Cy7-conjugated anti-CD11b (all from Biolegend, San Diego, CA, USA) at a concentration of 0.5 μg/mL for 30 min at 4 °C. The corresponding fluorophore-conjugated isotype controls were used for the gating of the positive-stained cells. Immunophenotypic analysis of 5000–10,000 cells of each sample was performed using FACSCanto II flow cytometer (BD Biosciences). The flow cytometry data were analyzed using Flowjo software (Tree Star, Ashland, OR, USA, ver. 10.0.7). Both assays were performed three times with duplicated samples.