Thin films of poly(ethylene 2,6-naphthalate) (PEN) were isothermally crystallized at 190 °C after being melted at 300 °C. Morphological observation by transmission electron microscopy (TEM) showed the spherulitic texture in the films. Selected-area electron diffraction (SAED) indicated that the crystals in the films are the a form, as expected from our thermal condition for crystallization. The SAED pattern from the untilted specimen was characterized by the fairly intense reflection ring accompanied by other weak rings, and this intense ring was indexed as 010. A series of SAED patterns, which were obtained from the same specimen area tilted at various angles in the TEM column, suggested that the crystallites are oriented with their (001) planes being preferentially parallel to the film surface. Subsequently, a set of the dark-field images of the two-dimensional spherulite taken by using two different parts of the 010 reflection ring revealed that most of the crystallites in such a spherulite are oriented with their (010) planes being parallel in its radial direction. In addition, the spherulites in small pieces (0.05–0.08 mm thick) of PEN, which had been crystallized under the same thermal condition as above, were determined to be negatively birefringent by polarizing light microscopy.