Coexpression and colocalization of CysLT1 and CysLT2 receptors by human MCs. Cytofluorographic analyses (A,B) showing expression of CysLT1 and CysLT2 receptor proteins by cord blood–derived hMCs and LAD2 cells. Cells were permeabilized for staining with the indicated anti–C-terminal Abs (A) or stained without permeabilization (B) with a polyclonal Ab against the third extracellular domain of CysLT1 (RB34) or with a monoclonal anti-CysLT2 (1B3) against the N-terminal 18 amino acids. The indicated blocking peptides were used to demonstrate specificity. (C) mBMMCs from the indicated genotypes were stained with RB34 with or without the corresponding blocking peptide. Shaded tracings in panels A to C are the isotype controls. Results in each panel are representative of 3 to 5 experiments for each. (D) Confocal images of LAD2 cells stained with directly labeled RB34 (Alexa Fluor 488) and anti–C-terminal CysLT2 Abs. Note staining of plasma membrane (blue arrows), nuclear envelope (white arrows), and nuclear inclusions (black arrows) and colocalization in each location. Results were similar regardless of which combinations of Abs were used. Results are from 1 experiment representative of 3 performed. Results with indirectly labeled Abs were similar (n = 2, results not shown). See “Immunostains and confocal imaging” for complete image acquisition information.
