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Fig. 4. Analysis of the mini-Plg/CD222 interaction. (A) Plg fragments were coated (10 µg ml^-1) on 96-well immunosorbent plates, which were blocked with 1% BSA. Then, the wells were incubated with purified CD222 for 4 hours at 4°C in BB. After washing twice with BB, bound CD222 was eluted using SDS-PAGE sample buffer and visualized by western blotting using the anti-CD222 mAb MEM-238. (B) Effect of CD222 expression on cell binding to mini-Plg. CD222^-/- mouse fibroblasts expressing human CD222 or human CD87 were allowed to attach to 96-well plates coated with Plg, mini-Plg or K1-3. The number of cells displayed is the number of cells attached to wells coated with the indicated proteins after subtraction of the cell binding to BSA. (C) Determination of the binding affinity of Plg fragments to CD222 by surface plasmon resonance. Purified CD222 was immobilized on a Biacore chip. The equilibrium dissociation constant K_d was calculated from the ratio of dissociation rate constant k_d and association rate constant k_a. Standard deviation was obtained from three measurements.

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