The widely used concentrations for stimulation of adhesion are TNF-α 10 ng/ml or LPS 1 μg/ml. However, it is not always the optimal concentrations for very good response/adhesion. You need to test a range of concentrations by yourselves to get the optimal concentrations for stimulation of your samples/tissue culture ( HUVEC etc). The protocol is attached as pdf. Here is the description of testing by flow cytometry.

Leukocyte to HUVEC adhesion assay (flow cytometry)

The adhesion of leukocytes to HUVEC was quantified by a flow cytometry assay allowing the evaluation of the adhesion of the three major leukocyte populations (granulocytes, monocytes and lymphocytes) to HUVEC monolayer under static conditions. With this method whole blood samples were first labeled with a mouse anti-human CD45 mAb directly-conjugated to a fluorochrome. CD45 antigen (also known as Leukocyte Common Antigen) is present on all leukocytes (lymphocytes, monocytes and granulocytes), but is absent on erythrocytes and platelets. The fluorescently-labeled leukocytes were then incubated with HUVEC monolayers (co-culture), and non-adherent leukocytes were subsequently removed. The remaining bound leukocytes were then visualized and quantified by using the characteristic morphologic and fluorescent parameters of the instrument. Blood samples (BC or FB) were divided into 100 μl aliquots in 5 ml steryle polystyrene round-bottom tubes (Falcon-Corning, cat. no. 352054) and stained with 2.5 μl of pre-titrated APC-H7-conjugated CD45 mAb for 15 min on ice in the dark. After incubation blood samples were treated with 3 ml of a lysis buffer containing (g/L) NH4Cl (8.248), KHCO3 (1.0) and EDTA (0.0368) in order to remove erythrocytes. Samples were incubated at room temperature for 5 min and gently vortexed. Samples were then centrifuged at 600g for 5 min, supernatants were removed and cells were washed one time with 1 ml steryle RPMI medium. Finally leukocyte samples were resuspended in 1 ml steryle RPMI medium for the subsequent co-culture. In some experiments neutrophils isolated from BC were used instead of total leukocytes in order to assess level of their adhesion to HUVECs, but the staining procedure was the same. Only confluent HUVEC monolayers were used during the co-culture with leukocytes. To this end, HUVEC maintained as above described, were plated in a 12-well plate at a density of 0.2 × 106 /ml in the medium. When cells reached 80–90% confluency, they were left unstimulated (medium alone) or stimulated with TNF-α for 24 h and LPS overnight. The concentrations of stimuli were chosen based on their ability to give optimal response on the basis of previous experiments (you have to test this in your own lab using a range of TNfa and LPS concentrations).

Before starting co-culture, HUVEC monolayers were carefully washed two times with sterile PBS; finally, 1 × 106 CD45-labeled leukocytes (1 ml of cell suspension) was added. In each experiment of co-culture, unstimulated and stimulated HUVECs alone (without leukocytes or neutrophils), as well as CD45-labeled leukocytes or isolated neutrophils were included as internal controls in order to make discrimination between this two populations during analysis. Samples were incubated for 30 min at 37 °C. After, incubation the medium was removed in order to eliminate nonadherent leukocytes, and HUVEC monolayers were carefully washed three times with sterile PBS. Finally, HUVECs with adhering leukocytes or isolated granulocytes were detached by using a non-enzymatic solution, cell suspensions were centrifuged at 600g for 5 min at room temperature and supernatants were removed. The pellets were resuspended in 350 μl PBS and kept on ice until flow cytometric acquisition.

The optimum concentration I found was 5 μg/ml of LPS