Endothelial cell lines express markers and are assumed to exhibit other endothelial cell responses. We investigated E-selectin expression from human umbilical vein endothelial cells, the spontaneously transformed ECV304 line and the hybrid line EA.hy926 by flow cytometry and immunofluorescence, mRNA and soluble E-selectin release. In cells exposed to tumour necrosis factor α (TNF-α) and interleukin-1β (IL-1β), median (range) percentage of E-selectin-positive HUVECs increased from 1.6(0.9–6.2)% to 91.4(83.0–96.1)%, (P=0.001) using flow cytometry. In contrast, E-selectin expression by ECV304 and EA.hy926 cell lines was 100-fold lower. E-selectin mRNA was detectable after 2h, maximal at 6h in HUVECs and undetectable in EA.hy926 and ECV304 cell lines after exposure to TNF-α/IL-1β. sE-selectin accumulation increased (P=0.004) in HUVECs only. Neutrophil adherence to ECV304 and EA.hy926 cells was poor compared to HUVECs (P=0.004). The cell lines ECV304 and EA.hy926 do not exhibit normal endothelium expression of E-selectin, and may not be appropriate for studies of adhesion.