Human monocytes comprise three distinct subsets, defined by their relative expression of CD14 and CD16. These subsets appear to have different functional roles within homeostasis and inflammation, but little is known about the manner in which they interact with macro-and microvascular endothelial cells, a key enabling component for the fulfillment of their functional roles. In the present study, we examined the locomotory behavior of the three major human monocyte subsets over human endothelial monolayers subjected to physiologically relevant levels of shear flow in vitro. Each subset was shown to preferentially perform different types of locomotory behavior in a resting state. A long-range crawling behavior, similar to the “patrolling” behavior of murine Ly6C− monocytes, was observed in CD14+ CD16− and CD14 dim CD16+ monocytes, but not in CD14+ CD16+ monocytes. CD14 dim CD16+ and CD14+ CD16+ monocytes showed a preference for adhering to microvascular over macrovascular endothelium, whereas CD14+ CD16− monocytes showed the opposite. Transendothelial migration was not observed in CD14 dim CD16+ monocytes during the 30-min observation period. Long-range crawling behavior in CD14 dim CD16+ monocytes was abrogated by blockade of ICAM1, VCAM1, or CX 3 CL1, in contrast with CD14+ CD16− monocytes, which only required ICAM1 for this behavior. These studies indicate the existence of subtype-specific human monocyte migratory behavior patterns with distinct adhesion molecule dependence, which may assist in elucidating their physiological function and relevance to disease.