Near-field scanning optical microscopy (NSOM) is used to characterize nanoscale topographic and fluorescence features in thermally annealed films of the conjugated polymer polyfluorene. Thin films of polyfluorenes with either two hexyl (1), octyl (2), or dodecyl (3) alkyl groups at the 9 position were studied. Annealed films were made by holding the films above their respective liquid crystalline phase transition temperature and then rapidly cooling the films. Upon annealing, the films show large spectroscopic and morphological changes. The emission spectra films of 1 show a large increase in emission at wavelengths greater than 500, while films of 2 and 3 show small or no change in the long wavelength emission. Polarized NSOM images of all three films show that the films organize into highly ordered nanoscale domains. The order in the films is found to be largest in the polymer with the shortest alkyl chains growing progressively less ordered with increasing chain length. Films of 3 have domains on the order of 15 nm, while films of 1 and 2 have domains 25-30 nm in size. The domains in films of 1 have additional translational order as they align into larger ribbonlike polymer structures. NSOM imaging at two wavelengths reveals that intra- and interpolymer emitting species are found nearly uniformly throughout all three films. Small insoluble clusters that remain in the annealed films show no contrast in the polarization or wavelength images. The spectroscopy and NSOM together show that close packing of polymer chains in films of 1 can provide highly ordered films but only at the expense of increased excimer emission. The dioctyl polymer (2) has an ideal alkyl chain length to be able to achieve high molecular order while maintaining a minimum of interpolymer interactions. Films of 3 with the longest alkyl substituent show poor polymer order while maintaining a substantial component of interpolymer emission.