Key methodological variables in endothelial-lined flow chamber studies. Flexibility in flow chamber design has resulted in a range of different endothelial-lined flow chamber setups. Listed here are key variables between those studies: (1) Flow chamber dimensions: to achieve laminar flow, a height/width ratio of <0.2 is recommended. (2) Presence/absence of RBCs: RBCs are required for platelet margination and may influence the efficacy of certain antithrombotic agents, like dipyridamole. (3) Culturing of ECs under static or shear stress conditions: ECs change phenotype upon sensing mechanical stress; hence, culturing under shear stress is advised. (4) Presence/absence of Ca²⁺ and Mg²⁺: physiological Ca²⁺ and Mg²⁺ levels are required for integrin function and for coagulation to take place. (5) Different EC sources: distinctive endothelial heterogeneity throughout the vasculature illustrates the importance of using an EC source appropriate to the research question. In addition, the expression level of several proteins that are involved in thrombus formation is affected by the passage number. (6) Confluency of EC layer: should be reported as a lack of confluency may trigger thrombus formation, because of exposure of subendothelial matrix components to the blood. Additional clarification is provided in the text. To enable comparison between studies, we encourage reporting the variables specified in this figure.