Subcellular localization of wild-type and mutant SMPD3 in MDCK cells. Tagged version of wild-type and mutant SMPD3 (G247S and D358G) were transfected, and their subcellular localization was determined by indirect fluorescence microscopy. More than 70% of stained cells within each sample showed similar staining as depicted in the panels. Magnification: 20×/0.75 NA Plan Apo objective of samples in Vectashield mounting medium for dluorescence with DAPI, H-1200 (Vector Laboratories, Burlingame, CA) stained with Alexa Fluor 488 goat antimouse IgG H+L (Invitrogen). Images were acquired using a Retiga 200R digital camera (QImaging, Surrey, BC) and HIS-Elements Ar 2.30 software (Nikon) and processed using Adobe Photoshop 7.0 (Adobe Systems, San Jose, CA).