Fig. 3. (A) Top, confocal xy-section through center of a cluster of living HEK CaR cells incubated with an extracellular fluorescent marker (fluorescein-dextran; 10,000 MW) illustrating limited aqueous spaces between cells. Bottom, `side-view' profile (yz) reconstructed from stack of confocal z-sections (each 1 µm thick) through the region indicated by the line shown in the top image (line width=3 pixels or 1.5 µm). (B) Co-immunostaining of CaR and PMCA in HEK CaR cell clusters. Top left, Alexa 488 staining of CaR (red pseudocolor); middle, CY5 labeling of PMCA (green pseudocolor); top right (merged image), yellow color depicts areas of overlap. Bottom panels show control staining for Alexa 488 + CaR peptide block (see Materials and Methods for details) and Cy5 secondary antibody alone.