Quantitative analysis of planar cell polarity. (a) Myosin II is planar polarized in the epidermis during elongation of the Drosophila embryo. Myosin II (red) localizes to vertical interfaces between anterior and posterior cells and Par3 (green) localizes to horizontal interfaces. Anterior is to the left and ventral is down in this image and in (b). (b) All cell interfaces in the image (red channel from (a)) were manually outlined in blue in order to quantify the orientation and mean fluorescence intensity of each interface.(c) The red channel in (a) and the blue lines in (b) were used to quantify the distribution of myosin II. Cell interfaces were grouped by orientation into 15° intervals. The absolute mean fluorescence intensity was quantified for each interval (left panel, sum of blue and red bars). Background was measured as the mean fluorescence intensity of the cytoplasm (left panel, red bars). Relative edge intensities were calculated using the raw data (center panel) or background correction (right panel). Values shown are relative to the mean fluorescence of horizontal interfaces (0-15°). The fold increase in myosin II at vertical interfaces (75-90°) in this example is 1.6 without background correction and 2.6 with background correction.