The fabrication and characterization of highly ordered thin films made from amphiphilic, regioregular polythiophene derivatives are described. Films of poly(3-(11-(2-tetrahydropyranyloxy)undecyl)thiophene (PTHPUDT) were prepared by the Langmuir-Blodgett technique. The amphiphilic nature of the polymer affords layer-by-layer deposition and the formation of multilayer films of head-to-head and tail-to-tail Y-type structure. X-ray diffraction studies indicate bilayer separations of approximately 30 A. Anisotropic optical absorption in the plane of the film indicates that the thiophene backbones are preferentially oriented along the dipping direction. Further, polarized light microscopy studies indicate that these films are highly birefringent and that the optical retardation is uniform over the entire film. Ellipsometry studies confirm the sizable magnitude of the birefringence. Optical second-harmonic generation studies of multilayer films provide information regarding both the thiophene orientation within the film and the anisotropic distribution of chromophores in the surface plane. Taken together, these data offer strong evidence of highly ordered films in which the hydrophobic polythiophene chains lie parallel to the substrate surface with their alkyl chains oriented normal to the surface, as dictated by the hydrophilic nature of the alkyl chain's terminal tetrahydropyran functional group. As such, these films offer the potential for elucidating the connection between polymer morphology and physical property in materials that are otherwise subject to a sufficiently complex distribution of morphologies that such a correspondence is precluded.