To control the molecular organization of octadecyl rhodamine B (RhB18), hybrid Langmuir-Blodgett monolayers containing clay minerals were prepared. The orientation and conformation of the RhB18 molecule were analyzed by polarized attenuated total reflection Fourier transform infrared (ATR-FTIR) spectroscopy and fluorescence spectroscopy. Monomers and dimers of RhB18 were observed by UV-Vis absorption spectroscopy. Infrared bands of RhB18 exhibited dichroism and shifted to higher or lower wavenumbers in films as a function of clay content. The main fluorescence band of RhB18 was more intense and shifted to lower wavelengths in films with higher clay content. In films without clay, the phenyl group of RhB18 is tilted relative to the xanthene group and both groups are tilted relative to the substrate. In films with clay, both the phenyl group and xanthene group are oriented parallel to the clay surface which is parallel to the substrate. The clay mineral surface induces a change in the conformation and orientation of the RhB18 chromophore.