P₄₀₀ protein is a concanavalin A (Con A)‐binding, 250–kilodalton glycoprotein characteristic of cerebellum. Extraction conditions for P₄₀₀ protein were investigated, and complete solubilization of P₄₀₀ protein from a submicrosomal fraction (P31 fraction) of mouse cerebellum was attained by the combination of 4% Zwittergent 3–14 and 4 M guanidinium chloride. The solubilized P400 protein was purified using Sepharose CL‐4B and Con A‐Sepharose chromatography. A monoclonal antibody (18A10) was prepared against P₄₀₀ protein. Endo‐β‐N‐acetylglucosaminidase F digestion of P₄₀₀ protein revealed that P₄₀₀ protein has a small number of asparagine‐linked oligosaccharide chains and that the epitope that is recognized by 18A10 monoclonal antibody is not on the asparagine‐linked oligosaccharide portion. Tissue distribution of P₄₀₀ protein was investigated by immunoblot analysis using 18A10 monoclonal antibody. P₄₀₀ protein was abundant in the cerebellum, but a very small amount of P400 protein or related antigen was also detected in other parts of the nervous system and in nonneural tissues. Immunohis‐tochemical studies indicated that P₄₀₀ protein was distributed abundantly in the soma, the dendritic arborization, and the axon of the Purkinje cell. No immunoreaction was observed in the other types of cells.