MANY technological and scientific applications have been proposed for Langmuir-Blodgett (LB) films1. These ordered arrays of oriented amphiphilic molecules may be useful as nonlinear optical systems2, as insulating or patterning layers in microelectronics3,4, as model systems for studies of two-dimensional phases5 and as molecular templates for protein crystallization6. The potential of LB films for these applications is sensitive to the details of their molecular packing; in particular, they require that the layers have a defect-free, periodic structure1. Here we present images from atomic force microscopy of domain boundaries between regions of different crystallographic orientation in LB multilayers. The regular lattice structure is preserved to within 1 nm of the grain boundaries, and the domains are oriented in a near-twinning arrangement. We also observe a periodic buckling superstructure along a particular lattice symmetry direction, with a wavelength of about 2 nm and an amplitude of ≤0.1 nm. The buckling was independent of surface pressure during deposition, dipping direction, number of layers deposited and nature of the substrate, and was stable over many hours. These departures from two-dimensional periodicity may have an important bearing on applications that rely on perfect crystallinity.