The Förster energy transfer between chromophores in several different types of Langmuir-Blodgett (LB) films exposed to aqueous conditions was examined. The donor and acceptor chromophores were either contained in a monolayer of a preformed copolymer incorporating amphiphilic side chains and hydrophilic spacer groups (dansyldihexadecylamine as the donor and 1,1′-dioctadecyl-3,3,3′,3′-tetramethylindocarbocyanine perchlorate as the acceptor) or grafted to a monolayer of such a copolymer (anthracene as the donor and 7-nitrobenz-2-oxa-1,3-diazole as the acceptor). The monolayers containing donor or acceptor chromophores were separated by neutral spacer layers of cadmium arachidate or copolymer material. It was found that the unattached chromophores diffused through the LB films during the experimental procedure and for several days afterwards. LB films containing chromophores that were grafted to copolymer and copolymer material spacer layers possessed high translayer structural order. There was evidence, however, that indicated that the structure relaxed with time.