This procedure describes the immunophenotypic characterization of menstrual blood-derived mesenchymal stem cells (MenSCs) by flow cytometry according to ISCT minimal criteria.
Harvest MenSCs using standard trypsinization (SOP-LB-003). Wash twice with PBS. Resuspend in FACS buffer at 1-5 × 10⁶ cells/mL.
Target cell count: 1 × 10⁶ cells per staining condition
Pass cells through 40 μm cell strainer to remove clumps. This is critical for accurate flow cytometry analysis.
Distribute 100 μL cell suspension per tube:
Add antibodies according to manufacturer's recommended dilution (typically 1:50 to 1:200). Mix gently.
| Marker | Fluorochrome | Typical Dilution |
|---|---|---|
| CD73 | APC | 1:100 |
| CD90 | FITC | 1:100 |
| CD105 | PE | 1:100 |
| CD34 | FITC | 1:50 |
| CD45 | PerCP | 1:100 |
| HLA-DR | PE | 1:100 |
Incubate in the dark at 4°C for 30 minutes. Protect from light throughout remaining steps.
Add 2 mL FACS buffer to each tube. Centrifuge at 300 × g for 5 minutes. Aspirate supernatant. Repeat wash once.
Resuspend cells in 300-500 μL FACS buffer. Add viability dye (7-AAD or DAPI) 5 minutes before analysis if required.
| Marker | ISCT Standard | Acceptance |
|---|---|---|
| CD73 | ≥95% | ≥90% |
| CD90 | ≥95% | ≥90% |
| CD105 | ≥95% | ≥90% |
| CD34 | ≤2% | ≤5% |
| CD45 | ≤2% | ≤5% |
| HLA-DR | ≤2% | ≤5% |
Generate report including:
| Version | Date | Description |
|---|---|---|
| 1.0 | 2026-02-21 | Initial release |