This work describes Langmuir-Blodgett (L-B) monolayer and multilayer assemblies constructed from a series of NLO-active azo-benzene derivatives possessing terminal moieties of variable dipole moment. The terminal groups are electron acceptors (acetyl, nitro, and cyano) and are connected to a common amphiphilic azo-benzene segment. Our experimental and theoretical results show that the interplay between two dominant non-covalent interactions within the assemblies, namely dipolar and pi-pi stacking interactions, dictate the packing density, structural order, as well as the electronic properties of the final films. L-B films of the acetyl derivative, which has the weakest total dipole across the azo-benzene chromophore, exhibits the highest packing density and the largest blue shift in the UV-visible absorption spectrum. This is rationalized by relatively strong pi-pi interactions between the azo-benzene chromophores overwhelming weak intermolecular dipole-dipole interactions. More importantly, the small internal dipole in the acetyl functional groups encourages packing in a configuration that lowers the overall energy and increases the packing density. In the case of the cyano and nitro derivatives, both L-B films show decrease in packing density and a weaker electronic coupling due to unfavorable overall dipole interaction that offsets the pi-pi interaction. We show that such unfavorable interactions lead to the formation of a staggered and loosely packed configuration. Our work demonstrates that a subtle difference in molecular structure can have a dramatic impact on aggregation, and consequently on the electronic and optical properties of nano-assemblies. This work demonstrates a way of controlling the formation of nanoscale structures at the molecular level through the control of noncovalent interactions. (C) 2007 Elsevier B.V. All rights reserved.