We performed SPR analysis using BIAcore T100 and performed the analysis of PD-L1-Ig, PD-L2-Ig binding to PD1-Ig as well as CD80-Ig binding to PD-L1-Ig and CTLA-4-Ig.
(A) Top row: Superimposed sensorgrams representative of PD-L1 and PD-L2 binding to PD-1-Ig chips; Lower row: Sensorgrams of PD-L1-Ig and CTLA-Ig binding to CD80-Ig chips. Proteins were injected for two minutes at a flow rate of 40 μl/min onto PD-1-Ig or CD80-Ig chips and allowed to dissociate for three more minutes. The data shown are representative of five separate experiments.
(B) Superimposed sensorgrams showing short (red) and long (black) injections of PD-L1 (top row) and PD-L2 (lower row) onto PD-1 chip respectively. Proteins at 10 μg/ml were injected for one (short) or seven minutes (long) at a flow rate of 10 μl/min onto the PD-1 chip. Sensorgrams were normalized in the Y axis and aligned in the X axis at the end of injection in order to align the dissociation phases up The data shown are representative of two separate experiments.