Differential binding of FX and Ac-LDL to SR-AI. (A-B) Increasing concentration of FX (0-40 µg/mL) (A) or increasing concentration of SR-AI inhibitor (Poly[I], polyclonal anti–SR-AI antibody, or Ac-LDL; 1-50 µg/mL) along with 1 µg/mL FX (B) were incubated in microtiter wells coated with hSR-AI (0.5 μg per well). Bound FX was probed using a peroxidase-labeled polyclonal anti-FX antibody and revealed by chromogenic conversion of tetramethylbenzidine. For the negative control, hSR-AI was omitted during the coating (○ in panel A). Data represent the mean ± SD (n = 3-7). (C-H) Confocal analysis of THP1-derived macrophages incubated (1 hour at 37°C) with 150 nM Alexa 488–labeled Ac-LDL (C-D), 10 μg/mL Alexa 488–labeled FX (E-F), or preincubated with FX prior to the addition of Alexa 488–labeled Ac-LDL (G-H). Polymerized actin was counterstained using Alexa 647–labeled phalloidin (C,E,G) or cells were immunostained for EEA-1 (D,F,H). Dotted lines define cell boundaries based on phalloidin staining. Arrows indicate area of colocalization. Z depth is 0.5 µm; bars represent 10 µm; objective, ×63. OD, optical density; PoAb, polyclonal antibody; Poly[I], polyinosinic acid.