Figure 2.

Effect of divalent cations on binding of locked αLβ2 or isolated I domains to immobilized ICAM-1. (A) Binding of K562 transfectants expressing αLβ2 containing wild-type (WT) or locked I domains to immobilized ICAM-1 was determined in 20 mM Tris⋅HCl (pH 7.5), 150 mM NaCl supplemented with 1 mM Mg²⁺ and Ca²⁺, 1 mM Mg²⁺, 1 mM Mn²⁺, 5 mM EDTA, or in medium containing Ca²⁺ and Mg²⁺ in the presence of the activating mAb CBR LFA-1/2 at 10 μg/ml as indicated. Numbers in parentheses are clone numbers of the K562 stable transfectants. (B) Effect of divalent cations on binding to ICAM-1 of K562 transfectants expressing isolated I domains. Binding was performed in Hepes/NaCl/glucose/BSA (20 mM Hepes, pH 7.5/140 mM NaCl/2 mg/ml glucose/1% BSA) supplemented with 1 mM EDTA, 1 mM Mg²⁺, or 1 mM Mn²⁺. Results are mean ± SD of triplicate samples and are representative of at least three experiments; some error bars are too small to be visible.