Antibody Conjugation to Cell Surface Receptors

Here, we describe the covalent modification of cell surfaces with antibodies through bioconjugation to the self-labeling SNAP-tag enzyme. We apply this approach to the reprogramming of T cell signaling through the universal chimeric antigen receptor (CAR), "SNAP-CAR." Universal CARs are a highly programmable class of engineered receptors that interact with co-administered "adaptor" antibodies to recognize one or more antigens of interest to trigger receptor signaling. Universal CARs have promise for use in cell therapeutics and as research tools. SNAP-CAR covalently attaches to adaptor antibodies containing a benzyl guanine (BG) motif. The protocols presented here include methods for SNAP-CAR T cell and antibody adaptor generation using gamma retroviral transduction and BG-NHS ester conjugation, respectively. The chapter also describes methods to evaluate cell surface bioconjugate formation, including quantification of BG motifs on antibodies and co-incubation experiments to assay for antigen-directed SNAP-CAR T cell functions of target cell killing and SNAP-CAR T cell activation.