We raised an anti-glioma monoclonal antibody, named G-22, that specifically recognizes a human glioma-associated surface antigen. Proven to be useful for target imaging of malignant gliomas after radioisotope labeling and cerebrospinal fluid diagnosis by enzyme-linked immunospecific assay, G-22 was found to immunoprecipitate an 85-kDa glycoprotein of the human glioma U-251MG cell. We purified this antigen by G-22-coupled cyanogenbromide-activated Sepharose affinity chromatography, and sequence analysis demonstrated that the 54 amino acid residues were identical to positions 55-108 of human CD44. The results show that the smallest spliced form (85 kDa) of CD44 is strongly expressed in glioma cells.