We propose a novel technique for characterizing interfacial structures in vertically aligned carbon nanofibers (CNFs) utilizing scanning transmission electron microscopy (STEM). In this technique, vertically aligned CNFs are selectively grown using plasma-enhanced chemical vapor deposition (PECVD), on a substrate comprising a narrow strip (width ∼100 nm) formed by focused ion beam. Using STEM, we obtain images of nanostructures of CNFs having diameters as small as 10 nm, while focusing on the interfacial region near the nanofiber base. Stacked graphite sheets parallel to the substrate are observed near the base of these CNFs.