Figure 1. SunTag-based clustering of αCD3/CD28 scFv (SBCS). (A, B) Schematic of the SBCS strategy for stimulation and expansion of T cells. (A) αCD3scFv or αCD28scFv fused with 12 tandem copies of the GCN4 tag was used to recruit αCD3/CD28 scFv fused with αGCN4scFv, forming 13×αCD3scFv or 13×αCD28scFv, respectively. (B) 13×αCD3scFv and 13×αCD28scFv bind to the TCR/CD3 complex and the CD28 molecule, respectively, to activate resting T cells. (C) Schematic drawing of the vectors used for recombinant protein expression. (D, E) SDS-PAGE analysis of the purified recombinant proteins of (C). All recombinant proteins were expressed by transient transfection of HEK293FT cells. Lanes 1, 2, and 3 of (D) represent αCD3scFv-4×GCN4, αCD3scFv-12×GCN4, and αGCN4scFv-αCD3scFv, respectively. Lanes 1 and 2 of (E) represent αCD28scFv-12×GCN4 and αGCN4scFv-αCD28scFv, respectively.