Three slices in total from the top,
middle and bottom of each cell z-stack were chosen for
quantification of colocalization with Manders coefficient.
Thresholds were adjusted manually to eliminate background
fluorescence signals. If necessary, a region of interest (ROI)
was drawn to image to exclude other cells or bacteria outside
the cell from the analysis. For each cell weighted arithmetic
mean of coefficients from three slices were calculated. Statistical
significances of colocalizations were calculated during
analysis using Costes algorithm, and only coincidence probabilities
of P ¼ 1.00 was taken into account.