Storage Temperature Stability of Therapeutic Cell Types
Therapeutic cells are "living drugs," and their viability and potency can change quickly if storage temperatures drift outside validated ranges. For most cell therapies, the safest long-term strategy is true cryogenic storage—kept below the glass transition (approximately –135°C for aqueous systems) to minimize damaging events like devitrification and ice recrystallization.[1,2,3,4]
Temperature Stability Thresholds
| Storage Condition | Temperature Range | Clinical Application |
|---|---|---|
| Cryogenic Storage | ≤ –150°C | Long-term stability for most cryopreserved therapeutic cells. Minimizes ice recrystallization. |
| Glass Transition | ~–135°C | Critical threshold where ice structure changes become possible. Above this, devitrification risk increases.[1,2,3,4] |
| Ultra-Low Freezer | –80°C | Not true cryogenic. Limited duration only; gradual viability loss without specific validation.[5] |
| Refrigerated | 2–8°C | Short-term hold. Metabolism slows but continues. Typically hours to 1–3 days depending on cell type. |
| Ambient | 20–25°C | Short window only. Cell stress accelerates. Generally hours-scale for most therapies. |
1. CAR-T Cells
Chimeric Antigen Receptor T-Cell ProductsStorage Requirements (Long-Term)
Commercial CAR-T products are typically labeled for storage in vapor-phase liquid nitrogen at very low temperatures (commonly ≤ –150°C, sometimes ≤ –120°C depending on product).[6,7,8,9,10,11]
Temperature Excursion Thresholds
Above ~–135°C: Crossing the glass transition threshold enables damaging ice structure changes.[1,2]
Dry ice (–78°C): Far warmer than cryogenic; limited transfer scenarios only if validated. Not equivalent to LN₂ storage.[5]
2–8°C (Fresh/Non-Cryopreserved): Short shelf-life requiring stability data support:[6,12]
Reduced post-thaw viability; shifts in phenotype or activation state; potency drift (killing capacity, cytokine profiles); increased lot-to-lot variability after similar handling.
CAR-T requires validated cold chain behavior: defined excursion limits, continuous monitoring, documented investigations when limits are exceeded.
2. Hematopoietic Stem/Progenitor Cells
HSC/HPC ProductsStorage Requirements
Long-Term (Cryopreserved): ≤ –150°C for banking, retention, and consistent post-thaw performance. FDA guidance addresses validated processes and ongoing quality control.[17]
Short-Term (Fresh Products):
- Refrigerated (2–8°C): Commonly used for ~48 hours, sometimes up to ~72 hours with validation
- Tran et al. (2024): refrigeration at 4°C better preserves attributes vs. room temperature[18]
- Jansen et al.: viable CD34+ cells retained during 4°C storage[19]
- Antonenas et al.: controlled cold storage reduces deterioration[20]
Lower viable CD34⁺ recovery; reduced colony-forming units (CFU); declining engraftment performance with prolonged non-ideal holds.
Clear split approach: (1) Cryogenic for banked or delayed products; (2) Refrigerated workflow only when infusion timing is predictable and tightly managed.
3. Mesenchymal Stromal Cells
MSC ProductsStorage Requirements
Long-Term (Cryopreserved): ≤ –150°C for consistency and banking
Short-Term Holds: Solution-dependent stability profile
- 4°C / 2–8°C: MSC survival varies 1–2 days in appropriate media; can collapse rapidly in simple saline-like buffers
- Nofianti et al.: adipose-derived MSC viability differed substantially between culture medium vs. normal saline, demonstrating formulation-dependent stability[21]
Critical Factor: Post-thaw time in cryoprotectant (DMSO) can be the limiting factor even when temperature is controlled. Many workflows require rapid dilution/removal and quick administration.
Delayed apoptosis after thaw; functional potency drift (immunomodulation, differentiation potential); phenotypic changes from stress and rewarming.
4. Induced Pluripotent Stem Cells
iPSC ProductsStorage Requirements
Cryopreserved: ≤ –150°C for banks and long-term stability. LN₂ storage is the default because conventional cryoprotectant systems can be thermally unstable at –80°C:[22,23]
- Yuan et al. (2016): long-term storage requires LN₂; –80°C enables ice recrystallization and progressive viability loss[22]
- –80°C storage considered short interim step only, not long-term unless validated
- 2–8°C refrigerated shelf life not standard practice
- Post-thaw: require rapid recovery into controlled culture conditions
Poor recovery/attachment after thaw; apoptosis and loss of colony-forming efficiency; drift in pluripotency markers and genomic stability.
Bank integrity: redundancy, documentation, and strict cryogenic continuity.
Regulatory Compliance Requirements
Quality expectations across CAR-T, HPC, MSC, and iPSC programs consistently include:
- Defined storage temperature ranges with validated stability claims
- Continuous monitoring (freezers, LN₂ systems, shippers)
- Documented excursion management: event details, duration, maximum temperature, impact assessment, disposition decision[7]
- Traceability and chain-of-custody for patient-specific products
- Release/lot acceptance criteria including viability and potency. FDA guidance: minimum acceptable viability specification generally 70% for somatic cellular therapies[12]
Summary: Temperature Control Ladder
| Temperature | Stability Profile | Application |
|---|---|---|
| ≤ –150°C | Best long-term stability | CAR-T, HPC, MSC, iPSC (cryopreserved) |
| ~–135°C | Glass transition threshold | Above this: increased risk |
| –80°C | Limited/validated only | Not "forever storage" |
| 2–8°C | Short hold window | Hours to ~1–3 days (cell type dependent) |
| 20–25°C | Shortest window | Hours-scale for most therapies |
Clinical Conclusion: Temperature excursions alter living product integrity. Modern biostorage requires validated cold chain execution: redundant systems, continuous monitoring, documented excursion response, and quality documentation meeting regulatory inspection standards.