CST, 93513S, CAR (G4S Linker) Antigen Density Quantitation Kit (PE)

Protocol
Available protocols: Flow Cytometry (Live)
Specificity / Sensitivity
G4S Linker (E7O2V) Rabbit Monoclonal Antibody (PE Conjugate) recognizes exogenously expressed levels of scFv-based CARs containing a G4S linker.
Species Reactivity: All Species Expected
Background
The stoichiometric relationship between surface expressed scFv-based chimeric antigen receptors (CARs) and their cognate antigens expressed on target cells is a critical determinant of the efficacy of CAR-engineered immune cells. As such, accurate measurement of CAR density on the cell surface is of paramount importance to assess how changes to CAR structure and cell engineering protocols affect CAR expression (1-15). The poly-Glycine-Serine (G4S) linker is a type of flexible, unstructured synthetic peptide linker sequence often leveraged to connect the variable heavy (VH) domain and variable light (VL) domain of single-chain variable fragments (scFvs) and CARs that utilize an extracellular domain scFv for target antigen recognition. The linker itself consists of a core pentapeptide sequence, Gly-Gly-Gly-Gly-Ser, that is repeated and commonly found as either a 15-mer (G4S) or 20-mer (G4S) within scFv-based CARs and scFv fragments. The linker sequence length plays a role in controlling scFv stability and the noncovalent association between the VH and VL domains (16,17).