The neural cell adhesion molecule (NCAM) is thought to have an important role in cell-cell interactions during development. To better understand NCAM function, we studied the adhesion of mouse N2A neuroblastoma cells and Chinese hamster ovary cells to different forms of NCAM using a quantitative centrifugal cell adhesion assay that measures the rate of cell removal from experimental substrates. Embryonic brain NCAM is highly polysialylated and contains both 180- and 140-kDa polypeptide isoforms, whereas embryonic retinal NCAM is less highly polysialylated and contains primarily the 140-kDa isoform. For both forms, cell adhesion to substrate-immobilized NCAM was temperature dependent, cation independent, and time dependent. Cell adhesion to NCAM substrates was not directly affected by drugs inhibiting cytoskeletal function or cellular metabolism, suggesting that NCAM function does not depend critically on cytoskeletal function or metabolic activity. Cell adhesion to retinal NCAM was blocked by anti-NCAM antibodies, and adhesion was increased by neuraminidase treatment of both types of NCAM. Adhesion to brain NCAM was effectively blocked by anti-NCAM antibodies only after neuraminidase treatment, suggesting that these cells adhere to highly sialylated and less-sialylated NCAM by different mechanisms. We propose that multiple mechanisms of cell adhesion involving NCAM may exist in different tissues during development and that the state of polysialylation of NCAM is important in regulating the relative importance of these mechanisms.