Genome editing in resting CD4+ T cells is intrinsically challenging due to limited viability and poor editing efficiency. This historically limits the downstream functional assays, including pathway analyses of these primary T cells. In this case study, developed in conjunction with Lonza, we will learn how using a combination of cell culture conditions, Synthego’s Research sgRNA, and Lonza’s 4D-NucleofectorⓇ enabled Dr. Manuel Albanese and his team to achieve unprecedented knockout editing efficiencies and sustained viability in primary T cells. These studies demonstrated that high-efficiency CRISPR T cell editing can have implications for successfully developing CAR-T cell immunotherapies to treat cancer and viral infections. Dr. Albanese and Dr. Adrian Ruhle conducted this study in Dr. Oliver Keppler’s lab at the Max von Pettenkofer Institute and Gene Center in Munich, Germany.

Learn how the authors achieved the following through a combination of optimal culture conditions and gene editing protocols:

  • High and consistent knockout efficiencies in resting primary human CD4+ T cells
  • Consistent single-gene knockout efficiency enabled experiments studying multigene knockouts
  • Ability to perform downstream functional studies due to sustained high viability