Description
The NanoSparkTM EVEN-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 EVEN-T protein complex allows for sterile filtration and easy removal of the activating reagent via standard media exchange or simple centrifugal cell washing. The percentage of CD4+ T cells can be increased by adding increasing amounts of EVEN-T to the culture media, effectively modulating the CD4+:CD8+ ratio during the expansion step. The functional dose range is media-specific and can be tuned to your platform needs.
NanoSparkTM EVEN-T: Breakthrough in T Cell Ratio Control
NanoSparkTM EVEN-T is first in class – there are no other T cell activator products that allow researchers to control the CD4+:CD8+ T cell ratio during expansion. 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 EVEN-T Soluble T Cell Activator Work?
The NanoSparkTM EVEN-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. EVEN-T is formulated with a safe, proprietary additive that enables shifting of the CD4+:CD8+ T cell ratio throughout the expansion.
The NanoSparkTM EVEN-T Soluble T Cell Activator activates resting human CD3+ T cells originating from purified T lymphocytes or patient peripheral blood mononuclear cells (PBMCs).
Inquire About Nanotein’s EVEN-T Soluble Activator
Need help getting started? Want to benefit most from your T cell activation and expansion? Contact the Nanotein team today to learn more.
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Convenient
Soluble, magnetic bead-free, protein-based, and ready-to-use.
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High Performance
Proprietary CD4+:CD8+ shifting technology with excellent total T cell expansion.
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Reliable
Employs anti-CD3/CD28 antibodies and a proprietary additive to modulate the CD4+:CD8+ ratio in a dose-dependent manner.
Documentation & Protocols
Consult the data sheets and supporting documentation for usage instructions and additional details, or explore the recommended protocols, which include suggested catalog numbers for cytokine supplements and media.
Data Sheets
EVEN-T Cell Data Sheet 2024 (IL-7 & IL-15)
The NanoSparkTM EVEN-T Soluble T Cell Activator is used in vitro to induce activation, modulate CD4+/CD8+ T cell ratio, and CAR gene delivery uptake T cells. 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 EVEN-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.
Certificate of Analysis (CoA)
Lot-specific EVEN-T CoA will be shipped with the product. Contact customer service if additional copies are required.
Nanotein Example EVEN-T CoA
Safety Data Sheet (SDS)
Carefully read product SDS before use.
NanoSparkTM EVEN-T SDS
Using IL-2 for 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.
Combining IL-7 and IL-15 for T cell expansion
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 EVEN-T Protocol and Data Sheet.
Tips for Best Results Using the NanoSparkTM EVEN-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 EVEN-T Soluble T Cell Activator for every mL of cell suspension (e.g., 1,000 µL of Activator for 100 mL of cell suspension) based on the media used per Figure 1.
- 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 EVEN-T Soluble T Cell Activator to the cell culture before adding the viral vector. If using electroporation, then electroporate 72 hours after activator addition.
Data & Research
Enhanced CD4+ T Cell Expansion Using NanoSparkTM EVEN-T Soluble T-Cell Activator
Increasing amounts of EVEN-T shifts the CD4+ :CD8+ ratio towards CD4+s with media-dependent differences in functional windows. NanoSparkTM EVEN-T Soluble T-Cell Activator was added to CD3+ negatively selected human peripheral blood T cells cultured in CellGenix’s TCM, Lonza’s T-Vivo, or FujiFilm’s Prime-XV serum-free media supplemented with IL-7 and IL-15 and compared to NanoSparkTM STEM-T Soluble T-Cell Activator. Cells were expanded for 10 days and analyzed on a flow cytometer on days 0, 7, and 10. Cells were labeled with CD4+ , CD8+ , CD45RA, CCR7, and CD95 fluorescent antibodies (Schmueck-Henneresse et al 2017). Expansion with NanoSparkTM EVEN-T Soluble T-Cell Activator modulates the expansion to shift the expanded T cell profile towards CD4+ T cells in a dose-dependent manner, effectively allowing the user to select a CD4+ :CD8+ T cell ratio at the outset of expansion.
Quality & Safety
The underlying NanoSparkTM platform has undergone biocompatibility testing when formulated with antibodies specific to our T cell activator, STEM-T, which we consider a significant derisking step for the platform as a whole. The details are as follows:
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:
