Migration of human melanoma (MV3) cells (i) is based on a coordinated formation and release of focal adhesion contacts mediated by integrin receptor molecules, (ii) requires the activity of the Na⁺/H⁺ exchanger NHE1 and (iii) depends on extracellular pH (pH_e). At focal adhesion sites, a local pH created by protons extruded by the NHE1 together with protons provided by the extacellular solution might affect the strength of the integrin/extracellular matrix bond and thereby modulate cell migration. We measured pH in very close proximity to the plasma membrane (pH_em). The outer leaflet of the plasma membrane was labeled with the phosphoethanolamine fluorescein DHPE, the glycocalyx with the fluorescein-conjugated wheat germ agglutinin (WGA). Labeled cells were continuously superfused with prewarmed HEPES-buffered Ringer solutions of various pH_e. pH_em de- or increased as pH_e was de- or increased. Cells created a proton gradient with the H⁺ concentration being higher at the leading edge than at the rear end. This gradient decreased as the H⁺ concentration of the superfusion solution increased. pH_em of cells initially treated with the NHE1 inhibitor cariporide (HOE642) decreased by up to 0.1 units as soon as HOE642 was removed. Thus, DHPE was incorporated only in the outer leaflet of the membrane. We conclude that NHE1 clearly contributes to the cell surface pH. The more alkaline pH_em at the rear end may facilitate the local release of focal adhesion contacts.