Moving your stem cell therapy from research to the clinic and beyond
Advancements in stem cell therapy continue to shed light on how stem cells may be used in various clinical applications, which include cell replacement therapies, immunomodulatory therapies, and many others. Whether you’re developing an induced pluripotent stem cell therapy or adult stem cell therapy, Thermo Fisher Scientific has the solutions to advance your research to the clinic and beyond. Advancing your stem cell therapy research to clinical treatment requires careful material selection because the quality of starting materials significantly impacts the properties of your final stem cell therapy product. We offer high-quality products for a variety of stem cell types.
iPSC workflow from isolation to characterization
Cell Therapy Systems iPSC Reagents
We offer solutions for every stage of your cell therapy development process. We have an extensive selection of high -quality research use only products to support your early cell therapy development. As you move toward clinical and commercial applications, our GMP-manufactured (21 CFR Part 820) Cell Therapy Systems (CTS) products offer specific cell and gene therapy intended use statements, extensive safety testing and proactive regulatory documentation.
Isolation and reprogramming
The first step in induced pluripotent stem cell (iPSC)–derived therapy generation is isolation and cultivation of the donor’s somatic cells. Many different cell types can be reprogrammed including dermal fibroblasts (e.g., from a skin biopsy) and a variety of blood-derived cells, like PBMCs, CD34+ cells, and T cells. The cell type used for reprogramming is often determined by availability of donor tissue as well as the ultimate purpose of the iPSC being generated.
Successful generation of iPSCs can depend on many factors including the age, health and disease status of the donor, the viability and proliferation rate of the isolated somatic cells, and the cell type. It is important to choose a reprogramming system which will help to overcome some of these potential challenges.
Sendai virus–based reprogramming
Sendai virus–based CytoTune reprogramming kits are one of the most versatile and robust commercially available reprogramming systems and can be used to reprogram a wide range of cell types and origins. Additionally, the CTS CytoTune-iPS 2.1 Sendai Reprogramming Kit is the first off-the-shelf reprogramming system designed for clinical and translational research.
Dynabeads Isolation kits
Gibco cell culture media
Cell Therapy Systems (CTS)—solutions designed to enable clinical and commercial GMP cell and gene therapy manufacturing
Cell engineering and gene transfer
Patient-derived iPSCs offer exciting potential in both cell therapy and in vitro disease modeling by enabling access to cell populations that are otherwise unavailable from living donors. With the recent discovery of site-specific gene editing, this true power is fast approaching. iPSCs can be genetically altered at a specific locus using genome engineering tools such as CRISPR or TALEN. In addition to genome editing tools we offer viral transduction solutions, lipid-based transfection reagents and electroporation tools. The Invitrogen Neon Transfection System delivers DNA, RNA, and protein into cells while avoiding the challenges faced by transfection reagents and viral methods. It is possible to achieve greater than 80% knock out of a specific locus for iPSCs by optimizing electroporation parameters and cell density with the Neon system.
Electroporation
Genome editing
Lipid-based transfection
Expansion, banking, and recovery
Considerations for banking PSCs
Generation of a robust PSC bank can be an important precursor to downstream cell therapy applications. Developed as a ready-to-use, xeno-free ancillary reagent, CTS PSC Cryomedium can be used to freeze PSCs, enabling large quantities of cells to be safely placed into long-term cryopreservation.
Recovery of cryopreserved PSCs
Careful handling of cryopreserved PSCs is critical to support post-thaw recovery while minimizing cell death. To reduce stress from osmotic shock when thawing PSCs in cryomedium, fresh culture medium should be added dropwise. Addition of RevitaCell Supplement post-thaw will further enhance PSC attachment and recovery.
PSC growth and expansion
When expanding PSCs in routine culture, maintenance of pluripotency, trilineage differentiation potential and karyotypic stability are essential for success in downstream cell therapy applications. CTS Essential 8 Medium was designed to provide a regulatory compliant expansion medium with components not directly derived from animals while maintaining these crucial PSC properties. Specialized culture vessels, such as Nunc Triple Flasks or Nunc Cell Factory Systems can facilitate large-scale expansion of adherent PSC cultures.
Banking and recovery
Growth and expansion
Differentiation
Induced pluripotent stem cells can generate virtually unlimited numbers of differentiated cell types, including neurons, cardiomyocytes, and potentially any other cell type in the body. These differentiated cells can then be used in a range of applications. Whether for basic research, drug discovery, or future therapeutic applications, stem cell differentiation requires standardized culture methods to ensure reproducible and reliable results. Gibco media, supplements, and substrates provide you with an easy-to-use, flexible set of tools for targeted differentiation to your desired cell lineage. Our differentiation portfolio simplifies your workflow and provides you with more control, allowing for faster, more efficient systems.
To view the complete differentiation portfolio, go to thermofisher.com/differentiation.
Characterization and release testing
Appropriate iPSC therapy product characterization remains central to the successful development of safe and efficacious cell therapies. Regulatory agencies require release testing to confirm the identity, purity, potency and safety of cell-based products. Thorough characterization and release testing of master cell banks is essential for successful development of stem cell therapies. A typical characterization workflow includes assessing pluripotency, genomic stability, and safety. Typical required safety assays include sterility, mycoplasma, endotoxin, and adventitious viral testing. More in-depth safety assessments may include HLA and cancer hot spot testing. See the ICH guidelines for validation of analytical procedures.
- PluriTest-Compatible PrimeView Assays for Pluripotency Confirmation
- TaqMan hPSC Scorecard Assay
- KaryoStat Assays for Assessing Genomic Stability of PSC Lines
- Oncomine Comprehensive Assay v3M
- AmpFLSTR Identifiler Direct PCR Amplification Kit
- MycoSEQ Mycoplasma Detection Kit
- LinkSēq HLA Typing Kits
- Flow Cytometry
Isolation and reprogramming
The first step in induced pluripotent stem cell (iPSC)–derived therapy generation is isolation and cultivation of the donor’s somatic cells. Many different cell types can be reprogrammed including dermal fibroblasts (e.g., from a skin biopsy) and a variety of blood-derived cells, like PBMCs, CD34+ cells, and T cells. The cell type used for reprogramming is often determined by availability of donor tissue as well as the ultimate purpose of the iPSC being generated.
Successful generation of iPSCs can depend on many factors including the age, health and disease status of the donor, the viability and proliferation rate of the isolated somatic cells, and the cell type. It is important to choose a reprogramming system which will help to overcome some of these potential challenges.
Sendai virus–based reprogramming
Sendai virus–based CytoTune reprogramming kits are one of the most versatile and robust commercially available reprogramming systems and can be used to reprogram a wide range of cell types and origins. Additionally, the CTS CytoTune-iPS 2.1 Sendai Reprogramming Kit is the first off-the-shelf reprogramming system designed for clinical and translational research.
Dynabeads Isolation kits
Gibco cell culture media
Cell Therapy Systems (CTS)—solutions designed to enable clinical and commercial GMP cell and gene therapy manufacturing
Cell engineering and gene transfer
Patient-derived iPSCs offer exciting potential in both cell therapy and in vitro disease modeling by enabling access to cell populations that are otherwise unavailable from living donors. With the recent discovery of site-specific gene editing, this true power is fast approaching. iPSCs can be genetically altered at a specific locus using genome engineering tools such as CRISPR or TALEN. In addition to genome editing tools we offer viral transduction solutions, lipid-based transfection reagents and electroporation tools. The Invitrogen Neon Transfection System delivers DNA, RNA, and protein into cells while avoiding the challenges faced by transfection reagents and viral methods. It is possible to achieve greater than 80% knock out of a specific locus for iPSCs by optimizing electroporation parameters and cell density with the Neon system.
Electroporation
Genome editing
Lipid-based transfection
Expansion, banking, and recovery
Considerations for banking PSCs
Generation of a robust PSC bank can be an important precursor to downstream cell therapy applications. Developed as a ready-to-use, xeno-free ancillary reagent, CTS PSC Cryomedium can be used to freeze PSCs, enabling large quantities of cells to be safely placed into long-term cryopreservation.
Recovery of cryopreserved PSCs
Careful handling of cryopreserved PSCs is critical to support post-thaw recovery while minimizing cell death. To reduce stress from osmotic shock when thawing PSCs in cryomedium, fresh culture medium should be added dropwise. Addition of RevitaCell Supplement post-thaw will further enhance PSC attachment and recovery.
PSC growth and expansion
When expanding PSCs in routine culture, maintenance of pluripotency, trilineage differentiation potential and karyotypic stability are essential for success in downstream cell therapy applications. CTS Essential 8 Medium was designed to provide a regulatory compliant expansion medium with components not directly derived from animals while maintaining these crucial PSC properties. Specialized culture vessels, such as Nunc Triple Flasks or Nunc Cell Factory Systems can facilitate large-scale expansion of adherent PSC cultures.
Banking and recovery
Growth and expansion
Differentiation
Induced pluripotent stem cells can generate virtually unlimited numbers of differentiated cell types, including neurons, cardiomyocytes, and potentially any other cell type in the body. These differentiated cells can then be used in a range of applications. Whether for basic research, drug discovery, or future therapeutic applications, stem cell differentiation requires standardized culture methods to ensure reproducible and reliable results. Gibco media, supplements, and substrates provide you with an easy-to-use, flexible set of tools for targeted differentiation to your desired cell lineage. Our differentiation portfolio simplifies your workflow and provides you with more control, allowing for faster, more efficient systems.
To view the complete differentiation portfolio, go to thermofisher.com/differentiation.
Characterization and release testing
Appropriate iPSC therapy product characterization remains central to the successful development of safe and efficacious cell therapies. Regulatory agencies require release testing to confirm the identity, purity, potency and safety of cell-based products. Thorough characterization and release testing of master cell banks is essential for successful development of stem cell therapies. A typical characterization workflow includes assessing pluripotency, genomic stability, and safety. Typical required safety assays include sterility, mycoplasma, endotoxin, and adventitious viral testing. More in-depth safety assessments may include HLA and cancer hot spot testing. See the ICH guidelines for validation of analytical procedures.
- PluriTest-Compatible PrimeView Assays for Pluripotency Confirmation
- TaqMan hPSC Scorecard Assay
- KaryoStat Assays for Assessing Genomic Stability of PSC Lines
- Oncomine Comprehensive Assay v3M
- AmpFLSTR Identifiler Direct PCR Amplification Kit
- MycoSEQ Mycoplasma Detection Kit
- LinkSēq HLA Typing Kits
- Flow Cytometry
Adult stem cell therapy workflow from isolation to characterization
We offer solutions for every stage of your cell therapy development process. We have an extensive selection of high -quality research use only products to support your early cell therapy development. As you move toward clinical and commercial applications, our GMP-manufactured (21 CFR Part 820) Cell Therapy Systems (CTS) products offer specific cell and gene therapy intended use statements, extensive safety testing and proactive regulatory documentation.
Cell isolation and culture
Hematopoietic stem cells (HSCs) and mesenchymal stromal cells (MSCs, also known as mesenchymal stem cells) are readily available from a variety of tissue sources and show potential to address many unmet medical needs including blood disorders and cancers (HSCs), tissue reconstruction and graft-versus-host-disease (MSCs).
Cell Therapy Systems—solutions designed to enable clinical and commercial GMP cell and gene therapy manufacturing
Considerations for HSC harvest and expansion
HSCs can be harvested from umbilical cord blood (CB), bone marrow (BM) and mobilized peripheral blood (mPB), and isolated using positive selection for CD34 surface marker expression. A major limitation of ex vivo expansion of harvested human hematopoietic stem-progenitor cells (HSPCs) is the rapid differentiation of HSPCs at the expense of primitive pluripotent HSCs. Most traditional and commercially available media systems result in differentiation of HSPCs and loss of HSCs capable of long-term bone marrow engraftment and immune reconstitution.
HSC isolation and expansion products
General tissue sample preparation and cell isolation
Considerations for MSC harvest and expansion
MSCs can be isolated from a wide range of tissues, the most common sources are bone marrow (BM-MSCs), umbilical cord blood and tissue (UB-MSCs and UC-MSCs respectively), and adipose tissue (adipose-derived or AD-MSCs). MSCs from bone marrow and cord blood can be isolated following density gradient centrifugation by directly seeding cells in culture medium. For tissue-based sources like adipose and umbilical cord (e.g., Wharton’s Jelly), enzymatic digestion is required prior to seeding cells onto cell culture treated vessels with culture media. If MSCs are isolated using serum-free media then the addition of 2.5% human AB serum during the initial culture step facilitates cell attachment and growth but is not required for subsequent passages. Since MSCs are characterized by their adherence to culture surface, the culture vessel is rinsed the following day with DPBS without calcium and without magnesium, and the medium is replaced to remove any non-adherent cells.
Following isolation the MSCs should be evaluated to ensure they meet the minimal criteria for define human MSCs are established by the Mesenchymal and Tissue Stem Cell Committee of the International Society for Cellular Therapy (ISCT) (Dominici, M. et al. (2006) Cytotherapy, Volume 8, Issue 4, 315–317):
Summary of criteria to define MSC
- MSC must be plastic-adherent when maintained in standard culture conditions
- MSC must express (≥95%+) CD105, CD73 and CD90, and lack expression (≤2%+) of CD45, CD34, CD14 or CD11b, CD79alpha or CD19 and HLA-DR surface molecules.
- MSC must differentiate to osteoblasts, adipocytes and chondroblasts in vitro
Cell engineering and gene transfer
Mesenchymal stem cells can be genetically altered using lipid-base transfection, electroporation, viral approaches or genome engineering tools such as CRISPR or TALEN. Methods may vary by cell type and application, so a broad array of gene transfer tools is available for your needs.
Electroporation
Lipid-based transfection
Cell counting
Prior to cell banking, a cell count should be performed. Total cell count should be determined in a manual, semi-automated or automated fashion.
- Countess II Automated Cell Counter
- EVOS Cell Imaging Systems
- CountBright Plus Absolute Counting Beads (flow cytometry based)
Once the cell number has been determined, the cells can be resuspended in final culture media system or prepared for cryopreservation.
Cryopreservation
Cryopreservation using defined, serum-free cryopreservation medium helps reduce the potential risk of adventitious agents or other animal-derived components being introduced to patients.
- CTS Dulbecco's Phosphate-Buffered Salines
- CTS PSC Cryomedium (recommended for HSCs)
- StemPro MSC SFM XF + 10% DMSO (recommended for MSCs)
- CTS Synth-a-Freeze Medium
Characterization and release testing
Appropriate HSC and MSC therapy product characterization remains central to the successful development of safe and efficacious cell therapies. Potency and safety assays are largely therapy specific.
Flow cytometry is used to evaluate cell viability and surface marker expression throughout the workflow, including immediately after isolation, post-expansion and after recovery from cryopreservation.
- Flow Cytometry instruments and reagents
A standard flow cytometry workflow can involve:
For HSC, a sample flow cytometry panel utilizes*:
- CD34 monoclonal antibody clone 581
- CD90 (Thy-1) monoclonal antibody
- CD45RA monoclonal antibody, clone MEM-56
- Human Hematopoietic Lineage Antibody Cocktail
- LIVE/DEAD Fixable Yellow stain
*All listed products are available with additional fluorophores and/or clones; LIVE/DEAD Fixable stains are available for multiple excitation/emission ranges
For MSC, a flow cytometry panel may involve the following:
- Antibodies:
- Negative markers (all same fluorophore):
- LIVE/DEAD Fixable Yellow stain
In addition to surface marker expression, per ISCT guidelines (Dominici, M. et al. (2006) Cytotherapy, Volume 8, Issue 4, 315–317) MSCs must maintain trilineage differentiation capacity. In addition to surface marker expression, per ISCT guidelines (Dominici, M. et al. (2006) Cytotherapy, Volume 8, Issue 4, 315–317) MSCs must maintain trilineage differentiation capacity.
Cell isolation and culture
Hematopoietic stem cells (HSCs) and mesenchymal stromal cells (MSCs, also known as mesenchymal stem cells) are readily available from a variety of tissue sources and show potential to address many unmet medical needs including blood disorders and cancers (HSCs), tissue reconstruction and graft-versus-host-disease (MSCs).
Cell Therapy Systems—solutions designed to enable clinical and commercial GMP cell and gene therapy manufacturing
Considerations for HSC harvest and expansion
HSCs can be harvested from umbilical cord blood (CB), bone marrow (BM) and mobilized peripheral blood (mPB), and isolated using positive selection for CD34 surface marker expression. A major limitation of ex vivo expansion of harvested human hematopoietic stem-progenitor cells (HSPCs) is the rapid differentiation of HSPCs at the expense of primitive pluripotent HSCs. Most traditional and commercially available media systems result in differentiation of HSPCs and loss of HSCs capable of long-term bone marrow engraftment and immune reconstitution.
HSC isolation and expansion products
General tissue sample preparation and cell isolation
Considerations for MSC harvest and expansion
MSCs can be isolated from a wide range of tissues, the most common sources are bone marrow (BM-MSCs), umbilical cord blood and tissue (UB-MSCs and UC-MSCs respectively), and adipose tissue (adipose-derived or AD-MSCs). MSCs from bone marrow and cord blood can be isolated following density gradient centrifugation by directly seeding cells in culture medium. For tissue-based sources like adipose and umbilical cord (e.g., Wharton’s Jelly), enzymatic digestion is required prior to seeding cells onto cell culture treated vessels with culture media. If MSCs are isolated using serum-free media then the addition of 2.5% human AB serum during the initial culture step facilitates cell attachment and growth but is not required for subsequent passages. Since MSCs are characterized by their adherence to culture surface, the culture vessel is rinsed the following day with DPBS without calcium and without magnesium, and the medium is replaced to remove any non-adherent cells.
Following isolation the MSCs should be evaluated to ensure they meet the minimal criteria for define human MSCs are established by the Mesenchymal and Tissue Stem Cell Committee of the International Society for Cellular Therapy (ISCT) (Dominici, M. et al. (2006) Cytotherapy, Volume 8, Issue 4, 315–317):
Summary of criteria to define MSC
- MSC must be plastic-adherent when maintained in standard culture conditions
- MSC must express (≥95%+) CD105, CD73 and CD90, and lack expression (≤2%+) of CD45, CD34, CD14 or CD11b, CD79alpha or CD19 and HLA-DR surface molecules.
- MSC must differentiate to osteoblasts, adipocytes and chondroblasts in vitro
Cell engineering and gene transfer
Mesenchymal stem cells can be genetically altered using lipid-base transfection, electroporation, viral approaches or genome engineering tools such as CRISPR or TALEN. Methods may vary by cell type and application, so a broad array of gene transfer tools is available for your needs.
Electroporation
Lipid-based transfection
Cell counting
Prior to cell banking, a cell count should be performed. Total cell count should be determined in a manual, semi-automated or automated fashion.
- Countess II Automated Cell Counter
- EVOS Cell Imaging Systems
- CountBright Plus Absolute Counting Beads (flow cytometry based)
Once the cell number has been determined, the cells can be resuspended in final culture media system or prepared for cryopreservation.
Cryopreservation
Cryopreservation using defined, serum-free cryopreservation medium helps reduce the potential risk of adventitious agents or other animal-derived components being introduced to patients.
- CTS Dulbecco's Phosphate-Buffered Salines
- CTS PSC Cryomedium (recommended for HSCs)
- StemPro MSC SFM XF + 10% DMSO (recommended for MSCs)
- CTS Synth-a-Freeze Medium
Characterization and release testing
Appropriate HSC and MSC therapy product characterization remains central to the successful development of safe and efficacious cell therapies. Potency and safety assays are largely therapy specific.
Flow cytometry is used to evaluate cell viability and surface marker expression throughout the workflow, including immediately after isolation, post-expansion and after recovery from cryopreservation.
- Flow Cytometry instruments and reagents
A standard flow cytometry workflow can involve:
For HSC, a sample flow cytometry panel utilizes*:
- CD34 monoclonal antibody clone 581
- CD90 (Thy-1) monoclonal antibody
- CD45RA monoclonal antibody, clone MEM-56
- Human Hematopoietic Lineage Antibody Cocktail
- LIVE/DEAD Fixable Yellow stain
*All listed products are available with additional fluorophores and/or clones; LIVE/DEAD Fixable stains are available for multiple excitation/emission ranges
For MSC, a flow cytometry panel may involve the following:
- Antibodies:
- Negative markers (all same fluorophore):
- LIVE/DEAD Fixable Yellow stain
In addition to surface marker expression, per ISCT guidelines (Dominici, M. et al. (2006) Cytotherapy, Volume 8, Issue 4, 315–317) MSCs must maintain trilineage differentiation capacity. In addition to surface marker expression, per ISCT guidelines (Dominici, M. et al. (2006) Cytotherapy, Volume 8, Issue 4, 315–317) MSCs must maintain trilineage differentiation capacity.
Intended uses of the products mentioned on the page vary.
For specific intended use statements, please refer to the product label.
