Description
The NanoSparkTM STEM-T Soluble T Cell Activator is a culture-ready reagent for in vitro activation and expansion of human T cells. The nanoscale of the NanoSparkTM STEM-T protein complex allows for sterile filtration and removal of excess reagent via either media exchange or simple centrifugal cell washing.
NanoSparkTM STEM-T: Breakthrough in T Cell Ratio Control
NanoSparkTM STEM-T is best in class, particularly for preserving stemlike CD8+ T cells important for higher efficacy. Competing products introduce clinical risks to the process by using synthetic ingredients that may contaminate T cell therapy if not painstakingly removed from the expanded culture.
The NanoSparkTM product line is modular and plug-and-play by design, using a proprietary self-assembly protein nanoparticle complex that is inherently soluble and can be filter sterilized.
How Does the NanoSparkTM STEM-T Soluble T Cell Activator Work?
The NanoSparkTM STEM-T Soluble T Cell Activator contains self-assembling protein nanoparticles with functional antibodies bound to the surface through high-affinity non-covalent interactions. These anti-CD3 and anti-CD28 antibodies are arranged in a proprietary biophysical combination on the NanoSparkTM platform to drive best-in-class performance that favors stemlike CD8+ T cell production.
The NanoSparkTM STEM-T Soluble T Cell Activator activates resting human CD3+ T cells originating from purified T lymphocytes or patient peripheral blood mononuclear cells (PBMCs).
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Protocols
The NanoSparkTM STEM-T Soluble T Cell Activator is used in vitro to induce activation and CAR gene delivery uptake in stemlike CD8+ cells via anti-CD3/CD28 pathways. It is intended for ex vivo activation and expansion of CD3+ T Lymphocytes (human resting T cells) from T cell isolates or peripheral blood mononuclear cells (PBMCs). The activator should be stored at -80 °C long-term. Once thawed, store at 4 °C protected from light for up to one month.
The NanoSparkTM STEM-T Soluble T Cell Activator is designed for use with a cytokine-supplemented T cell expansion medium. Suggested protocols have been developed for common cytokine-supplemented strategies. Optimization of the provided general protocol information may be necessary depending on your experimental objectives.
IL-2 Supplementation for Optimal T Cell Proliferation
Interleukin 2 (IL-2) is a potent general T cell growth factor essential for the long-term proliferation of activated T cells.
IL-2 should be added to culture media at 20 ng/mL final concentration. For detailed information, please refer to the IL-2 STEM-T Protocol and Data Sheet.
IL-7 and IL-15 for Enhancing T Cell Survival and Homeostasis
Interleukin 7 (IL-7) is a cytokine required in early T cell development that promotes T cell survival by up-regulating the expression of anti-apoptotic genes. Interleukin 15 (IL-15) is an inflammatory cytokine that supports the homeostasis of naive CD8+ T cells. Combining IL-7 and IL-15 in culture promotes the enrichment of self-renewing T cells like TSCM.
IL-7 and IL-15 should be added to culture media at 10 ng/mL final concentration of each. For detailed information, please refer to the IL-7/IL-15 STEM-T Protocol and Data Sheet.
For use in Wilson Wolf Sterile GRex6M cell culture vessels, please see IL-7/IL-15 GRex6M STEM-T Protocol and Data Sheet for details.
Tips for Best Results Using the NanoSparkTM STEM-T Soluble T Cell Activator
- Seed CD3+ T cells/PBMCs at 7 x 105 cells/cm2 in culture media.
- Add 5-10 µL of NanoSparkTM STEM-T Soluble T Cell Activator for every mL of cell suspension (e.g., 1,000 µL of Activator for 100 mL of cell suspension).
- Incubate cells at 37 °C and 5% CO2 in a humidified incubator.
- For cryopreserved cells, gently thaw cells and exchange them into culture media before starting.
- For optimal cell expansion and maintenance, ensure the activator is in culture media (conditioned or fresh) for at least 72 hours (removal is unnecessary for expansion).
- Be sure to add fresh cytokine-supplemented culture media every three days
If using transduction (optional), wait 24-72 hours after adding NanoSparkTM STEM-T Soluble T Cell Activator to the cell culture before adding the viral vector. If using electroporation, then electroporate 72 hours after activator addition.
Quality & Safety
Nanotein assessed the NanoSparkTM platform with adherence to rigorous ISO 10993 standards in preparation for GMP manufacturing. The composition of the NanoSparkTM reagent is protein-based, and consequently naturally biodegradable, which provides a significant safety advantage over magnetic bead-based and synthetic products. A reputable medical device CRO was commissioned to complete the in vitro & in vivo ISO 10993 biocompatibility testing. The tests were run at >20X the predicted possible carry-over amount, which resulted in no toxicity or adverse effects:
ISO 10993 Biocompatibility Testing |
Test |
Type of Test |
Result |
Product Tested |
ISO 10993-5 |
Cytotoxicity |
In vitro |
No Toxicity/Adverse Effect |
NanoSpark STEM-T |
ISO 10993-10 |
Sensitization |
In vivo: Guinea Pig |
No Toxicity/Adverse Effect |
NanoSpark STEM-T |
ISO 10993-10 |
Irritation/Intracutaneous Reactivity |
In vivo: Rabbits |
No Toxicity/Adverse Effect |
NanoSpark STEM-T |
ISO 10993-11 |
Acute Systemic Toxicity |
In vivo: Mice |
No Toxicity/Adverse Effect |
NanoSpark STEM-T |
ISO 10993-11 |
Pyrogenicity |
In vivo: Rabbits |
No Toxicity/Adverse Effect |
NanoSpark STEM-T |
ISO 10993-4 |
Hemocompatibility |
In vitro |
No Toxicity/Adverse Effect |
NanoSpark STEM-T |
ISO 10993-3 |
Genotoxicity Ames |
In vitro |
No Toxicity/Adverse Effect |
NanoSpark STEM-T |
ISO 10993-3 |
Genotoxicity Mouse Lymphoma Forward Mutation |
In vitro |
No Toxicity/Adverse Effect |
NanoSpark STEM-T |