Fig. 1

Myoferlin and TOM20 poorly colocalize. A-C Indirect immunofluorescence performed on Panc-1 and MiaPaCa-2 cell lines, using TOM20 rabbit monoclonal and myoferlin (D-11) mouse monoclonal antibodies (A) High magnification showing myoferlin proximity (white arrows) with TOM20 in Panc-1 and MiaPaCa-2 cell lines. Colocalized pixels are pointed with yellow arrows. Scale bars represent 5 μm. B Low magnification showing myoferlin and TOM20 localization within Panc-1 and MiaPaCa-2 cells. White arrows pointed at region with high TOM20 staining while myoferlin staining was low. Scale bar represents 8.89 μm in Panc-1 and 5 μm in MiaPaCa-2 cell lines. The confocal pictures were acquired with a high resolution LSM 880 microscope. C Quantification of colocalization between TOM20 and myoferlin using Manders’ method. M1 represents the proportion of TOM20 colocalizing with myoferlin, while M2 represents the proportion of myoferlin colocalizing with TOM20. For the positive control, two secondary antibodies carrying distinct fluorochromes (Alexa Fluor 488 and 546), recognized the same myoferlin rabbit polyclonal primary antibody (HPA). The experiment was performed as three biological replicates. Each dot represents one individual cell. The non-parametric test of Kruskal-Wallis was performed for statistical analyses. Results are presented as mean ± SD, ns: non-significant. D Western blot showing myoferlin abundance in mitochondrial fractions. Calreticulin was used as an ER marker, while TOM20 was used as a mitochondrial marker. GAPDH was used as a loading control. These western blots are representative of three biological replicates