Prokaryotic high-level expression and activity assay of extracellular domain of B7-2 (CD86) in vitro

Objective: To clone, express and investigate the costimulatory activity of B7-2(IgV + C), extracellular domain of B7-2(CD86). Methods: B7-2(IgV + C) gene was amplified from B7-2 cDNA by PCR and then cloned into an prokaryotic expression vector pGEX-4T-3. The engineered E. coli were used to express protein. After denaturation and renaturation, the protein was used to stimulate human T lymphocytes cooperated with anti-CD3 in vitro. ³ H-TdR incorporation was used to detect the activation of T cells. Results: The plasmid pGEX-4T-3/hB7-2(IgV + C) was reconstructed and expressed a M, 55 × 10³ fusion protein in E. coli at a level of 33% of the total cellular protein. The experiment in vitro suggested that the fusion protein activated human T lymphocyte. Conclusion: With the first signal, the recombinant protein B7-2(IgV + C) had costimulatory avtivity to activate T cells.