Cell surface glycoproteins of human tumor cell lines (melanomas and astrocytomas, and ovarian, bladder, stomach, cervical, laryngeal, and renal cancers) were studied by labelling with 1) neuraminidase-galactose oxidase-[3H]borohydride, 2) galactose oxidase-[3H]borohydride, and 3) dilute periodate-[3H]borohydride. The labeled components were examined by polyacrylamide gel electrophoresis and fluorography. Each tumor type had a distinctive pattern of labeled glycoproteins when the results from both procedures 1 and 2 were considered. Cell surface glycoproteins of malignant melanoma could not be labeled by procedure 2, whereas the other cell lines had at least two major glycoproteins that could be labeled by this method. Very similar profiles of melanoma glycoproteins were labeled by procedures 1 and 3. From these results the conclusion was reached that cell surface glycoproteins of melanomas are substituted with sialic acid so that their D-galactose and/or N-acetyl-D-galactosamine residues are available for oxidation by galactose oxidase only after neuraminidase treatment. An alternative explanation that these sugars are sterically accessible to galactose oxidase only after neuraminidase treatment also has to be considered. All melanoma lines studied were characterized by having two major cell surface glycoproteins with molecular weights of 110,000 and 90,000, respectively. Lines, however, varied considerably in their expression of other components. In particular, heterogeneity was shown in the expression of gp220, a component identified as fibronectin by immunoprecipitation with a specific antiserum, and in the expression of gp37/32, a pair of glycoproteins having the characteristics of la-like antigens. Of the other cell lines studied, astrocytomas most closely resembled melanoma in their glycoprotein profiles. The brain tumors, however, had two or three glycoproteins, including gp110, which could be labeled by galactose oxidase-[3H]borohydride without neuraminidase treatment.