iPSC Technology Reprogramming and QC
Induced pluripotent stem cells (iPSC) technology has provided new tools to improve drug discovery efforts from efficacy and toxicity testing to novel target identification and understanding disease mechanisms. iPSC are generated from primary human cellular material using reprogramming technologies that direct the differentiated primary cell into a pluripotent state. The process is carefully executed and controlled to ensure that the resulting iPSC line is viable and properly characterized. These iPSC can be developed into relevant disease models because:
- They have a human genetic background
- They can be differentiated into multiple cell types for further study
- The production process allows for extensive expansion to ensure sufficient, consistent study material.
Stage
Description
QC
1Primary Material
Primary tissue is received, thawed, expanded, banked and QC screened.
Primary tissue sources:
- Whole blood or isolated PMBC
- Skin biopsies or derived fibroblasts
- Glioma tissue
- Urine or derived epithelial cells
- Microbiology: Mycoplasma qPCR and Bacteria/Fungi Growth in Broth
- Viral Screen: HIV-1, HIV-2, HBV, HCV qPCR
- Identity: STR Profile, confirmation
- Karyotype: array CGH
2Reprogramming
The Sendai virus method is used to generate stable clones.
3Clone selection
Multiple clones are chosen, single cell cloned and tested.
- Confirm absence of Sendai reprogramming vector
- Morphology
- Growth and viability profile
- Sterility
4Clone Expansion and Banking
Selected clones are expanded to the desired cell number and banked.
5Post Bank QC
Extensive QC is done to ensure that the cell line is viable and properly characterized
- Growth & Viability: assessment of recovery post thaw
- Microbiology: Mycoplasma qPCR & Bacteria/Fungi Growth in Broth
- Viral Screen: HIV-1, HIV-2, HBV, HCV qPCR
- Identity: STR Profile, confirmation of genetic lesion
- Vector Clearance: Confirmation via qPCR
- Karyotype: array CGH & G-banding
- Phenotype: Flow cytometry
- Differentiation: EB Scorecard
6Quality Assurance
- Version controlled documents
- Internal audits
- Certificate of Analysis
- Approval of critical suppliers/service providers
- Independent Quality Assurance staff
