Deletion of <i>flotillin 1</i> does not alter clustering of APP in mouse embryonic fibroblasts.

<p><b>A.</b> Immunofluorescence staining for endogenous flotillin-1, which resides in flotillin microdomains at the plasma membrane, does not overlap with GFP-APPswe in images acquired with TIR illumination. Pearson's correlation coefficient calculated from 10 similar images is shown. <b>B.</b> GFP-APPswe colocalised extensively with antibody against clathrin heavy chain, detected by indirect immunofluorescence and TIR illumination. Pearson's correlation coefficient calculated from 10 similar images is shown. <b>C.</b> Consistent with previous reports, no significant overlap was observed between flotillins and clathrin, detected as in B. Pearson's correlation coefficient calculated from 10 similar images is shown. <b>D.</b> PALM during TIR illumination was used to determine the size of mEos2-APPswe clusters at the plasma membrane of MEFs. A representative image after particle detection and reconstruction with QuickPALM is shown. Fitted centroids from the PALM analysis are represented as a single pixel. The intensity of that pixel is proportional to the accuracy of the fitted centroid, and the intensities of centroids fitted to the same pixel are summed. E. Frequency distribution of cluster size for mEos2-APPswe at the plasma membrane, comparing control and <i>flotillin 1-/-</i> MEFs. At least 10 images were analysed for each genotype.