Anti-PECAM/NC internalization is inhibited in REN cells expressing isoforms that lack tyrosine 686. (A) Cells were incubated with FITC-labeled anti-PECAM/NCs at 37°C for 3 hours, washed, fixed, and counterstained with Texas red goat anti–mouse IgG to label noninternalized nanocarriers. Merged images show internalized anti-PECAM/NCs as single-labeled green particles (arrows) and surface-bound anti-PECAM/NCs as double-labeled yellow particles (arrowheads). (i) RhP. (ii) ΔCD. (iii) Δ11-16. (iv) Δ15-16. (v) Y663F. (vi) Y686F. Magnification bar represents 10 μm. (B) The percentage of internalization was calculated as mean plus or minus SD (n ≥ 25 cells; *P < .001).