Table 1 Quality controls performed at (a) the different steps of the culture and their impact on batch validation and quality controls performed at (b) the thawing of the cells and their impact on batch validation
From: Clinical-scale expansion of mesenchymal stromal cells: a large banking experience
Test | Culture step | Material | Method | Specifications | Release criteria for downstream manufacture |
|---|---|---|---|---|---|
(a) MSC quality controls | |||||
Cell count | P1, P2, Harvest | Cell suspension | Cell counting (Neubauer cell counting chamber) | None | Informative |
Cell viability | P1, P2, Harvest | Cell suspension | Trypan blue exclusion (Neubauer cell counting chamber) | None | Informative |
Morphology | P1, P2, Harvest | Cell suspension | Microscope observation | Fibroblastic | Release criteria for batch validation |
Aspect | P1, P2, Harvest | Cell suspension | Microscope observation | Cell suspension without aggregates | Release criteria for batch validation (harvest only) |
Sterility | P0, P2 and harvest | Culture supernatant | BactAlertR | Sterile | Release criteria for batch validation (P2 and harvest) |
Mycoplasma | P2 and harvest | Culture supernatant | Biochemical test (detection of mycoplasma enzymes by bioluminescence) | Absence of mycoplasma | Release criteria for batch validation |
Endotoxin | P2 and harvest | Culture supernatant | LAL (limulus amebocyte lysate) detection E.P. 2.6.14. | Endotoxin level <2.5 U.I./ml | Release criteria for batch validation |
Cell identity and purity (phenotype) | P2 and harvest | Cell suspension | Facs analysis | CD90Â >Â 70Â % CD105Â >Â 70Â % CD73Â >Â 70Â % CD14Â <Â 5Â % CD34Â <Â 5Â % CD45Â <Â 5Â % CD3Â <Â 1Â % | Release criteria for batch validation |
Karyotype | P2 and harvest | Cell culture | Chromosome analysis | Absence of abnormal chromosomal structure and/or number | Release criteria for batch validation |
Immunosuppressive properties | Harvest | Cell culture | MLR with stimulated PBMC-cell cycle analysis | >25Â % Inhibition of PBMC-induced proliferation (10/1 ratio) | Release criteria for batch validation |
(b) Post-thaw MSC quality controls | |||||
Cell count | Thawing | Cell suspension | Cell counting (Neubauer cell counting chamber) | Yes, cell number should be > × 10E6 and < × 10E6 cells/kg recipient According to the clinical protocol | Release criteria |
Viability | Thawing | Cell suspension | Trypan blue exclusion (Neubauer cell counting chamber) | 50Â % | Release criteria |
Sterility | Thawing | Thawed cell product | BactAlertR | Sterile | No, results are not known at the time of infusion. Cells are released according to sterility results at harvest |