Using BACS™ microbubble technology, the Human T Cell Selection, Activation and Expansion Kit was designed to gently isolate highly pure T cells and activate them for use in cell therapy workflows. In this positive selection workflow, T cells are isolated, activated, and expanded using Akadeum’s Selection, Activation & Expansion Microbubbles in combination with an antibody cocktail that targets CD3 and CD28. This highly scalable process yields large numbers of activated and transducible cells needed to support research and process development in T cell based therapies. The gentleness of flotation-based separation and activation results in fewer exhausted cells compared to legacy activation methods – better research and process development outcomes.
Greater activation and lower cellular exhaustion for healthier cells, enabling better therapies
Improved expansion of highly activated effector T cell populations.
Scalable: process billions of cells with the Alerion™ Microbubble Cell Separation System
Closed-system in-bag workflows available
GMP and GMP Grades available to support each stage of your cell therapy pipeline
Akadeum’s novel approach to positive selection for CAR-T cell production provides improvements over current industry standard methods, and the innate scalability of BACS™ microbubbles allows for rapid optimization of cell therapy workflows.
Using washed leukopak material or PBMCs as a starting material, T cells are labeled with biotinylated antibodies targeting CD3 and CD28. BACS™ Selection, Activation & Expansion Microbubbles are mixed into the sample where they bind to the labeled cells and gently float them to the surface. Unwanted cells can be removed from below the microbubble layer, and the positively selected T cells transferred to a culture vessel.
BACS™ are inherently inert and do not impact culture conditions. Over time and with cell division, cells gently settle to the bottom of the plate where they continue to expand until collection.
Additional Information:
Compatible with the Alerion™ Microbubble Cell Separation System
Antibody Cocktail (biotinylated): CD3 and CD28
Storage: 4°C
Cell Type: T Cells
Sample Species and Source: Human PBMC or platelet washed Leukopak material
Go from research scale to commercialization with microbubbles. Just use more or less to suit your research needs and keep performance maintained.
Experimental Overview:
T cells were positively selected from PBMC input material at 100 million total cells (5 mL tube), 1 billion total cells (a 150 mL bag), or 5 billion total cells (BioRise™ Consumable and Alerion™ Microbubble Cell Separation System) following the Human T Cell Selection, Activation, and Expansion protocol.
A) Estimated time based on protocol steps (mixing, incubation, separation) for separation completed in each vessel. As cell numbers increase the workflow time does not lengthen.
B) Comparison of cell viability results at day 1 post selection for cells separated in 5 mL tube, 150 mL bag, and BioRise™ Consumable. All protocols resulted in high cell viability.
C) Total cell expansion results comparing the BioRise™ Consumable selected cells to 5mL tube and 150mL bag vessel controls. Five million captured T Cells were seeded for expansion using G-Rex® 6M Well Plates and complete ExCellerate™ media supplemented with 2% human serum. Additionally, 5U/mL IL2, 5ng/mL IL7, and 5ng/mL IL15 was added on days 0, 4, and 7 of culture. No significant differences were observed in growth rates between the different isolation vessels.
The Right Products to Support Your Pipeline
The Human T Cell Selection, Activation and Expansion Kit is ideal for research through process development, with GMP and GMP versions available to support preclinical to commercialization of cell therapy workflows.
Data
More Activation, Less Exhaustion. Healthier Cells.
Optimize results while minimizing potential cellular stress or damage.
2 x 108 PBMCs were used as starting material for the Akadeum Human T Cell Selection, Activation, and Expansion kit workflow. Post selection, 5 x 106 T cells/well were seeded and cultured in G-Rex® 6M well plates (Wilson Wolf Manufacturing). Magnetic particles (following the manufacturer’s recommended protocol) were used for comparative T cell selection and were seeded similarly at 5 x 106 per well. Cells for control wells were isolated via Akadeum’s Human T Cell Leukopak Isolation Kit (negative selection) and seeded at 0.5 x 106 per mL in standard 24-well tissue culture plates. Complete ExCellerate™ Media (Bio-Techne), supplemented with 2% human serum (Access Biologicals) was used for all conditions. Additional IL-2, IL-7, and IL-15 cytokines (PeproTech), were added for Akadeum Human T Cell Selection, Activation, and Expansion kit wells while magnetics and control wells were supplemented with IL-2. Harvest timepoints for all conditions were on days 4, 7, and 11.
Resources
User Guide
For processing on the Alerion™ Microbubble Cell Separation System, please refer to:
App Notes
Publications & References
Video
Follow along with our expert-in-residence, Jason Ellis, PhD, as he demonstrates the protocol for using Akadeum’s Human T Cell Selection, Activation, and Expansion Kit with PBMCs and apheresis materials.
FAQs
Cell Therapy Products
What are the minimum and maximum cell numbers that can be processed with Akadeum’s kits?
It is not recommended to process under 10 million cells per sample. The maximum number of cells processed is dependent on the size of the vessel as each kit is very scalable.
What selection method is used for Akadeum’s kits?
Akadeum’s Human T Cell Leukopak Isolation Kit is the first ever negative selection kit that can meet the scale requirements for cell therapy. The Human T Cell Depletion Kit also uses negative selection to deplete endogenous CD3+ T cells. The Human T Cell Selection, Activation, and Expansion Kit uses positive selection.
What is meant by washed leukopak material?
Washed leukopak material refers to a leukopak that has been processed to reduce platelets, exchange buffer, and concentrate the cells. This can be done manually via centrifugation or by using commercially available cell washing instruments.
Can unwashed leukopaks be used with Akadeum’s kits?
Yes, Akadeum has successfully processed raw leukopaks. Please email techsupport@akadeum.com to learn more.
Can Akadeum’s kits be used with cryopreserved samples?
Yes, Akadeum’s kits are designed to work with both fresh and cryopreserved samples.
Are Akadeum’s kits for autologous or allogeneic workflows?
Akadeum’s kits can be integrated into both autologous and allogeneic workflows.
General Questions
Are Akadeum microbubbles hollow silica spheres?
Yes, Akadeum’s microbubbles are hollow silica spheres. Silica is a material that has extensive precedence for use in therapeutics and diagnostics.
Are antibodies directly attached to the microbubbles?
Only specific kits, such as our Dead Cell Removal (DCR) and RBC microbubble kits, use antibody-coated microbubbles. Most Akadeum microbubble products are not coated with antibodies; instead, a biotinylated antibody cocktail is provided to use with the streptavidin-coated microbubbles.
What is buoyancy-activated cell sorting (BACS)?
BACS is Akadeum’s proprietary cell separation technology that uses buoyant microbubbles to isolate specific cell populations utilizing only the power of gravity.
What are microbubbles?
Akadeum’s microbubbles are hollow silica spheres that are on average 12 microns.
How big are microbubbles?
Microbubbles average 12 um in diameter. Learn more about microbubbles technology.
How many cells can a microbubble lift?
Microbubbles can carry as many as four cells.
What is the Separation Buffer?
Ca2+, Mg2+ free Dulbecco’s PBS containing 2mM EDTA, and 0.5% bovine serum albumin (BSA). BSA must be biotin free to not interfere with kit performance.
After sitting in the refrigerator, all the microbubbles have separated out of solution. Is there something wrong?
No, this is normal due to the buoyant properties of microbubbles. The solution will need to be mixed well before every use. If processing multiple tubes, mixing between tubes is necessary.
I don’t have an aspirator/vacuum available in my lab, can I still use Akadeum microbubbles?
Aspiration is the best and easiest way to remove the microbubble layer after centrifugation. Pipetting off the bubble layer is not very effective. Vacuum pumps are available from a variety of vendors.
How are Akadeum’s kits shipped?
Kits are shipped on ice packs with overnight delivery.
I have additional technical questions, who should I contact?
Please email all technical questions to techsupport@akadeum.com or check out our chatbot!
