A comparison has been made of various methods for measuring binding of nerve growth factor (NGF) to PC12 cells in suspension, on plates, and by a combination of the two. Results indicated that the extensive washing in the plate binding assay removed some cell surface ligand, underestimated the fast receptor binding, and overestimated the proportion of internalized ligand. In addition, the binding and internalization by a nonresponding PC12 mutant cell line has been studied. The nonresponding mutants had fewer total NGF receptors (10-50%) than normal cells in any binding assay. However, when measured in the suspension assay, the mutant cells showed both fast and slow binding receptors, in proportion approximately equivalent to those found on native PC12 cells. The PC12 nonresponders in suspension were also found to internalize and degrade low levels of NGF, in proportion to their reduced receptor number. Different results concerning PC12 wild type and mutant cells that have been reported in the literature may be due to the particular binding assay protocol that was used.