Ez link maleimide peg11 biotin reagent

The ligands, NF-pCj1–HLA-DP5^ec and pCj1–HLA-DP5^ec, were biotinylated with the EZ-Link Maleimide-PEG₁₁-Biotin reagent (Thermo Fisher Scientific), according to the manufacturer’s instructions, and then immobilized in a flow cell containing a streptavidin sensor chip (Series S Sensor Chip SA; GE Healthcare), leading to ~1000 resonance units (RU). The SPR measurements were performed on a Biacore T200 (GE Healthcare) instrument by the injection of a series of analytes, TCR(SU3–3)^ec diluted with HBS-P buffer (GE Healthcare) to various concentrations (0.3125–10 μM), for 120 s at a flow rate of 10 μl min⁻¹ at 20°C, using HBS-P buffer. The final response curves were obtained by subtracting the curve obtained in the absence of the analyte. The equilibrium dissociation constant (K_D) values of NF-pCj1–HLA-DP5^ec·TCR(SU3–3)^ec and pCj1–HLA-DP5^ec·TCR(SU3–3)^ec were calculated with the BIAevaluation software (GE Healthcare).

The ligands, NF-pCj1–HLA-DP5^ec and pCj1–HLA-DP5^ec, were biotinylated with the EZ-Link Maleimide-PEG₁₁-Biotin reagent (Thermo Fisher Scientific), according to the manufacturer’s instructions, and then immobilized in a flow cell containing a streptavidin sensor chip (Series S Sensor Chip SA; GE Healthcare), leading to ~1000 resonance units (RU). The SPR measurements were performed on a Biacore T200 (GE Healthcare) instrument by the injection of a series of analytes, TCR(SU3–3)^ec diluted with HBS-P buffer (GE Healthcare) to various concentrations (0.3125–10 μM), for 120 s at a flow rate of 10 μl min⁻¹ at 20°C, using HBS-P buffer. The final response curves were obtained by subtracting the curve obtained in the absence of the analyte. The equilibrium dissociation constant (K_D) values of NF-pCj1–HLA-DP5^ec·TCR(SU3–3)^ec and pCj1–HLA-DP5^ec·TCR(SU3–3)^ec were calculated with the BIAevaluation software (GE Healthcare).