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Quantification of ICAM-1 adhesion.

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posted on 2010-06-28, 01:03 authored by Jaap D. van Buul, Jos van Rijssel, Floris P. J. van Alphen, Mark Hoogenboezem, Simon Tol, Kees A. Hoeben, Jan van Marle, Erik P. J. Mul, Peter L. Hordijk

(A) αICAM-1-antibody coated-beads were allowed to pre-adhere to TNF-α-stimulated endothelium for 5 minutes. Experiment was carried out as described in Materials and Methods. Briefly, low shear was introduced (0.25dyn/cm2) and flow was increased stepwise with 1.25dyn/cm2 at the time points, indicated with arrows. The final shear corresponded to 6.25dyn/cm2. The αICAM-1-antibody coated beads detached from the endothelium when shear increased. Pre-treatment of the endothelium with cytochalasin B (CytoB) or NSC-23766 (NSC) showed a steeper decline of detached beads upon increased shear flow. Experiment is done three times. Data are mean ± SEM. *p<0.05. (B) Experiment carried out as described under A. Pre-treatment of the endothelial cells with either blebbistatin (Blebbi) or cyclodextrin (β-CD) show a decrease in adhesion of the beads to the endothelium, already after the first increase in shear flow. Experiment is done three times in duplicate. Data are mean ± SEM. *p<0.01.


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