Highly efficient non-viral delivery of macromolecules to unactivated human T cells using microfluidic transfection

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Presentation Time: 11:30 AM - 12:45 PM
Poster Board Number: B923
Abstract ID: 5530

Presenting Author:
Manreet Chehal , Scientist at STEMCELL Technologies

Abstract:

Traditional methods for transfecting unactivated human T cells are limited in their delivery efficiency while maintaining cell viability, phenotype, and function. To address these challenges, we developed the CellPore™ Transfection System. This microfluidic platform creates transient disruptions in the plasma membrane via pressure-induced mechanical deformation, allowing for cytosolic delivery.

 

First, we developed a workflow to optimize delivery pressure to T cells using FITC-dextran. This optimized pressure was used to deliver mRNA for expression of eGFP (91.8%) and mCherry (92.9%), and Cas9 RNP complexes targeting B2M and TRAC, with knockout efficiencies of 74.0% MHC-I and 93.9% TCRαβ, respectively. Importantly, while electroporation of T cells resulted in increased expression of the activation marker CD69 (41.2%) and proinflammatory cytokines (77.7 pg/mL IL-2, 8.8 pg/mL IFNγ), CellPore™-transfected T cells maintained an unactivated phenotype (0.8% CD69+, 0.1 pg/ml IL-2 and 0.2 pg/ml IFNγ) and were capable of downstream activation/expansion via CD3, CD28, and CD2 cross-linking. Using this optimized workflow, efficient cytosolic delivery of FITC-dextran to human CD4 (97.4%), CD8 (96.6%) and regulatory (98.3%) T cells was also achieved.

 

The CellPore™ Transfection System enables robust and efficient delivery of cargo to T cells without impacting cell function in a simple workflow that can easily be integrated into research protocols.