FIG. 4.
Quantitative image analysis of a fixed adipocyte expressing EGFP/PLCδ1-PH (A) and labeled with the membrane-staining probe Alx594-ConA (B). (F) Plot of normalized fluorescence intensities (green line, EGFP fluorescence; red line, Alx594 signal) along the cell boundary (i.e., red lines in panels A and B). Pix. Int., pixel intensity. Blue line, baseline. (C and G) Ratiometric intensities (G) and corresponding ratiometric image (C) generated as described in the legend to Fig. 3. (D and E) Magnified views of the boxed regions (box shown only in panel B) in panels A and B, respectively. Peak EGFP fluorescence intensities corresponding to PtdIns(4,5)P2-rich membrane domains are labeled 1 to 9 in panels A and F. Green and red arrows in panel A indicate the starting and ending points of the line intensity profile (clockwise), respectively. Consequently, a portion of the cell boundary suspected of being broken is excluded from data analysis. Scale bar, 5 μm.