Project Summary/Abstract Cellular therapies, in which living cells are the drug administered to patients, have the potential to advance treatment for a variety of inherited and acquired diseases. In particular, chimeric antigen receptor (CAR)-T therapies have shown remarkable efficacy in certain hematological cancers. The first generation of approved CAR-T therapies rely on viral vectors for cellular reprogramming. Viral vectors have enabled the transduction of tough-to-transfect primary T cells but have several drawbacks related to their relatively lengthy manufacturing processes, immunogenicity, and potential for insertional mutagenesis. Furthermore, the field is trending towards more complex reprogramming methods, such as multiple gene edits via CRISPR technology, that are not compatible with typical packaging limits associated with viral approaches. Electroporation is a well-known method for delivery of DNA, RNA, and proteins into cells, that is recognized as the leading replacement for viral delivery. CyteQuest is dedicated to address the need for a flexible electroporation platform that can facilitate discovery and easily scale cell therapy manufacturing from the research level (low-volume) to the clinical level (high-volume). CyteQuest has developed a scalable electroporation platform to optimize transfection parameters and deliver cargo efficiently and reproducibly at high throughput. The platform incorporates a single use, continuous-flow fluidic system designed to integrate with automated cell processing approaches. The key objectives of this proposal are to: (1) design, construct, and test optimization instrument prototypes capable of multiplexed electroporation (up to parallel 8 channels) to enable rapid optimization of transfection parameters and use these instruments with academic and clinical collaborators to advance their cell therapy programs; and (2) design, construct and test manufacturing instrument prototypes capable of rapid delivery of cargo to mid and high volumes of cells to enable GMP-compatible cell manufacturing within a closed system. CyteQuest’s goal is to expand the availability of cellular therapy to treat a wider range of diseases and extend treatment to a much broader patient community.