Fig. 4.

CD9 is involved in optimal VCAM-1 binding and modulation of the integrin activation state. (A) L3055-12 and L3055-33 were stained with anti-CD9, anti-CD29, and CD49d. Histograms with broken and solid lines indicate isotype control and specific antibody, respectively. (B) The binding of soluble VCAM-1-Fc (10 μg/mL) by L3055-12 and L3055-33 was measured by flow cytometry. Competition experiments were performed to confirm the binding specificity by incubating neutralizing anti-VCAM-1 with VCAM-1-Fc for 30 min before being added to L3055-12 cells. Histograms with broken and solid lines indicate the binding to 41BB-Fc (negative control) and VCAM-1-Fc, respectively. The filled histogram indicates the binding to VCAM-1-Fc after the preincubation with anti-VCAM-1. (C) L3055-12 and L3055-33 were stained with anti-CD29 (clone MAR4, total CD29 expression, filled histograms) and anti-CD29 (clone HUTS21, activated form of CD29 expression, empty histograms) following incubation with 41BB-Fc (negative control) and VCAM-1-Fc. Histograms with a broken line indicate the isotype control.