Multilayer dye aggregation at the dye/TiO2 interface of dye-sensitized solar cells is probed via first principles calculations, using p-methyl red azo dye as an example. Our calculations suggest that the multilayer dye aggregates at the TiO2 surface can be stabilized by π…π stacking and hydrogen bond interactions. Compared with previous two-dimensional monolayer dye/TiO2 model, the multilayer dye aggregation model proposed in this study constructs a three-dimensional multilayer dye/TiO2 interfacial structure, and provides a better agreement between experimental and computational results in dye coverage and dye adsorption energy. In particular, a dimer forms by π…π stacking interactions between two neighboring azo molecules, while one of them chemisorbs on the TiO2 surface; a trimer may form by introducing one additional azo molecule on the dimer through a hydrogen bond between two carboxylic acid groups. Different forms of multilayer dye aggregates, either stabilized by π…π stacking or hydrogen bond, exhibit varied optical absorption spectra and electronic properties. Such variations could have a critical impact on the performance of dye sensitized solar cells.