Protocol for engineering primary human and mouse T cells using peptide-assisted genome editing

Efficient engineering of human and murine primary T cells without perturbation of cellular function is crucial for immunotherapy and immunology research. Here, we present a protocol for genome editing of primary T cells with minimal toxicity using peptide-assisted genome editing. We describe key steps required for primary T cell isolation and activation, CRISPR ribonucleoprotein (RNP) assembly and delivery mediated by peptide, and assessment of editing efficiency. This protocol maintains T cell proliferative capacity and functional potency. For complete details on the use and execution of this protocol, please refer to Zhang et al.¹.