Active T-Cell Immunoreceptor With Ig And ITIM Domains Protein (TIGIT)

T cell immunoreceptor with Ig and ITIM domains (TIGIT) is an inhibitory receptor expressed on several types of lymphocytes. TIGIT is expressed on NK cells and subsets of activated, memory and regulatory T cells, and particularly on follicular helper T cells within secondary lymphoid organs. Human TIGIT cDNA encodes 244 amino acids (aa) including a 21 aa signal sequence, a 120 aa extracellular region with a V-type Ig-like domain and two potential N-glycosylation site, a 21 aa transmembrane sequence, and an 82 aa cytoplasmic domain with an ITIM motif. A 170 aa variant diverges after aa 166. Lymphocyte Activation Gene 3 (LAG3) and TIGIT are two potential targets for cancer immunotherapy, thus a functional binding ELISA assay was conducted to detect the interaction of recombinant human TIGIT and recombinant human LAG3. Briefly, biotin-linked TIGIT were diluted serially in PBS, with 0.01% BSA (pH 7.4). Duplicate samples of 100 μl were then transferred to LAG3-coated microtiter wells and incubated for 1h at 37℃. Wells were washed with PBST 3 times and incubation with Streptavidin-HRP for 30min, then wells were aspirated and washed 5 times. With the addition of substrate solution, wells were incubated 15-25 minutes at 37 ℃. Finally, add 50 µl stop solution to the wells and read at 450 nm immediately. The binding activity of recombinant human TIGIT and recombinant human LAG3 was shown in Figure 1, the EC50 for this effect is 0.11 ug/mL.