Transient Expression of a Soluble and Secreted form of Heterodimeric T-Cell Receptor in Hek-293

Abstract

Biologically active human T-cell receptor (TCR) was expressed as a secreted recombinant protein in 293 cells. Correct α/ß chain pairing was facilitated by complementary charged peptides (leucine zipper) at the C terminus. Two plasmids, coding for the α and ß chains were constructed and prepared using commercially available purification kits. After optimizing transfection conditions for secretion of heterodimeric TCR in 12 well plates, transfections were performed in spinner flasks. The transfection efficiency was determined with a co-transfected ß-gal expression vector nd correlated with TCR levels as determined by ELISA. Product was isolated by affinity chromatography. Biological activity, i.e. ligand binding was tested. Transiently produced recombinant soluble TCR showed similar affinity to the ligand as receptors purified from cell membranes.