Layered perovskite La2CuO4+δ (LCO) thin films were epitaxially grown on SrTiO3 (STO) and LaSrAlO4 substrates by atomic-layer-by-layer molecular beam epitaxy. The lattice defects and residual strain in these films were investigated by means of transmission electron microscopy and electron energy loss spectroscopy. The LCO films showed a high epitaxial quality with flat interfaces and top surfaces. Misfit dislocations with Burgers vector a<010> and shear defects were frequently observed at or near the film/substrate interfaces and in the films, respectively. In one LCO film, grown on STO at the highest temperature, 700 °C, we observed a two layered structure with the top layer being rhombohedral CuLaO2. In addition, stacking faults were observed in the plane views of one LCO film grown on the STO substrate. The residual strains were evaluated from the associated splitting of Bragg reflection. Possible mechanisms of strain relaxations are discussed based on the observed defects.