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Fabrication and Use of Dry Macroporous Alginate Scaffolds for Viral Transduction of T Cells. | LitMetric

Fabrication and Use of Dry Macroporous Alginate Scaffolds for Viral Transduction of T Cells.

J Vis Exp

Joint Department of Biomedical Engineering, University of North Carolina at Chapel Hill and North Carolina State University; Comparative Medicine Institute, North Carolina State University; Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill;

Published: September 2022

Genetic engineering of T cells for CAR-T cell therapy has come to the forefront of cancer treatment over the last few years. CAR-T cells are produced by viral gene transfer into T cells. The current gold standard of viral gene transfer involves spinoculation of retronectin-coated plates, which is expensive and time-consuming. There is a significant need for efficient and cost-effective methods to generate CAR-T cells. Described here is a method for fabricating inexpensive, dry macroporous alginate scaffolds, known as Drydux scaffolds, that efficiently promote viral transduction of activated T cells. The scaffolds are designed to be used in place of gold standard spinoculation of retronectin-coated plates seeded with virus and simplify the process for transducing cells. Alginate is cross-linked with calcium-D-gluconate and frozen overnight to create the scaffolds. The frozen scaffolds are freeze-dried in a lyophilizer for 72 h to complete the formation of the dry macroporous scaffolds. The scaffolds mediate viral gene transfer when virus and activated T cells are seeded together on top of the scaffold to produce genetically modified cells. The scaffolds produce >85% primary T cell transduction, which is comparable to the transduction efficiency of spinoculation on retronectin-coated plates. These results demonstrate that dry macroporous alginate scaffolds serve as a cheaper and more convenient alternative to the conventional transduction method.

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Source
http://dx.doi.org/10.3791/64036DOI Listing

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