Syk was enriched at the leading edge upon β₂ integrin–mediated adhesion. Confocal microscopy images of human Syk detected by a mouse anti–human Syk mAb and a secondary Alexa 546–conjugated anti–mouse IgG Ab (red) or Syk-EGFP epifluorescence (green). Murine Syk was detected using a rabbit anti–mouse Syk Ab and an Alexa 546–conjugated anti–rabbit IgG Ab (red). F-actin was assessed using Alexa 488–conjugated phalloidin (green), EGFP-actin epifluorescence (green), or Alexa 633–conjugated phalloidin (red). (A) Upon induction of adhesion of untransfected dHL-60 cells (WT), EGFP-Syk– or EGFP-actin–transfected dHL-60 cells by fMLP (100 nM) on immobilized fibrinogen, F-actin and Syk were enriched at the leading edge (arrow). The merged images demonstrate colocalization (arrow, yellow). (B) Upon induction of adhesion of EGFP-Syk–transfected dHL-60 cells by fMLP (100 nM) on immobilized fibronectin, the enrichment of Syk at the leading edge was absent (arrowhead). (C) Upon induction of adhesion of untransfected dHL-60 cells by Mn²⁺ (0.2 mM) on immobilized fibrinogen, Syk was enriched at the leading edge (arrow), whereas TNFα (30 ng/mL) induced cell spreading without an obvious formation of a leading edge. (D) Upon induction of adhesion of murine CD18^+/+ PMNs on immobilized fibrinogen by fMLP (100 nM), Syk was enriched at the leading edge (arrow), whereas Syk was homogenously distributed in CD18^–/– PMNs, which showed a defect in cell polarization. Transmission images show the morphology of the corresponding cell (transmission). Scale bar equals 10 μm; n = 3.