We compared the effects of two anti-beta1 integrin activating antibodies, TS2/16 and AG89, on K562 cell adhesion to fibronectin. Though both antibodies effectively induced cell adhesion, the EC50 for AG89 was more than 200-fold higher than that for TS2/16. Scatchard analysis of the data from [125I]Fab fragment binding to the cells revealed that the TS2/16 epitope is exposed constitutively on all the beta1 integrin molecules, while only 3% of the beta1 integrins on resting K562 cells bear the AG89 epitope. Calculation of the actual number of each antibody bound to the cell during the cell adhesion assay revealed that induction of cell adhesion can be accomplished by binding much fewer AG89 molecules compared to TS2/16. Thus, AG89 and TS2/16 represent distinct classes of anti-integrin activating antibodies that show completely different binding characteristics as well as different activation effects on the integrin molecule upon binding.