A Laminar Flow-Based Assay to Study the Interactions Between Leukocytes and Endothelial Cells

Begin by placing a glass slide with an adherent cytokine-stimulated endothelial monolayer into a flow chamber.

These endothelial cells express surface adhesion molecules, such as selectins, and endothelium-bound migratory signals, like chemokines.

Introduce fluorescently labeled leukocytes to the flow chamber.

As leukocytes flow over the endothelial monolayer, they engage with activated endothelial cells.

Selectins on endothelial cells bind to their corresponding ligands on the leukocytes.

Selectin-ligand interactions are weak and transient, enabling leukocytes to roll along the endothelial monolayer under the force of a flowing medium.

Rolling allows leukocytes to interact with chemokines, activating integrins and promoting firm leukocyte arrest.

Subsequently, leukocytes spread across the endothelial monolayer, signifying their potential for transmigration, a crucial step in the immune response.

This assay helps to study the mechanism of adhesion, de-adhesion, and transmigration of leukocytes while perfusing them over endothelial cells.

Use one micromolar of cell-permeant green fluorescent nucleic acid stain to fluorescently label the leukocytes. After labeling, incubate the leuko