DePauw University 1 article published in JoVE Biology Serial Block-Face Scanning Electron Microscopy (SBF-SEM) of Biological Tissue Samples Justin A. Courson1, Paul T. Landry1, Thao Do1, Eric Spehlmann2, Pascal J. Lafontant2, Nimesh Patel1, Rolando E. Rumbaut3,4, Alan R. Burns1,4 1College of Optometry, University of Houston, 2Department of Biology, DePauw University, 3Center for Translational Research on Inflammatory Diseases (CTRID), Michael E. DeBakey Veterans Affairs Medical Center, 4Children’s Nutrition Center, Baylor College of Medicine This protocol outlines a routine method for using serial block-face scanning electron microscopy (SBF-SEM), a powerful 3D imaging technique. Successful application of SBF-SEM hinges on proper fixation and tissue staining techniques, as well as careful consideration of imaging settings. This protocol contains practical considerations for the entirety of this process.