Interaction of large unilamellar vesicle (LUV) of dipalmitoylphosphatidylcholine (DPPC) with ethanol was investigated by the excimer method developed by Yamazaki et al. (Yamazaki, M., M. Miyazu, and T. Asano. 1992. Biochim. Biophys. Acta. 1106:94-98) and the high-resolution electron cryomicroscope with a new cryostage (top-entry superfluid stage) (HiRECM) developed by Fujiyoshi, Y. et al. (Fujiyoshi, Y., T. Mizusaki, K. Morikawa, H. Aoki, H. Kihara, and Y. Harada. 1991. Ultramicroscopy. 38:241-251). The excimer method is based on the fact that the ratio of excimer to monomer fluorescence intensity (E/M) of pyrene PC is lowered in the membrane in the interdigitated gel structure (L beta I), because structural restriction of L beta I structure largely decreases collisions of pyrene rings of the pyrene PCs in the membrane. E/M of pyrene PC in DPPC LUV decreased largely at high concentrations of ethanol, which indicated the induction of L beta I structures in DPPC LUV. Frozen-hydrated DPPC LUVs in a vitreous ice were observed at 4K with HiRECM, and these images were characterized by a pair of concentric circles. The membrane thickness of DPPC LUV which was estimated from the distance between the two concentric lines decreased largely at high concentration of ethanol. The mean value of membrane thickness of the LUV in the absence of ethanol was 3.8 nm, while at 15% (w/v) ethanol was 3.0 nm. These values were almost same as those obtained from the electron density profile of DPPC MLV by the x-ray diffraction analysis in each structures, L beta' and L beta I structures, respectively. These results indicated directly the induction of L beta 1 structure in DPPC LUV at high concentration of ethanol.