The DS19 Friend erythroleukemia cells possess peripheral-type benzodiazepine recognition sites that are similar to those characterized in other tissues and cells. These sites recognize only specific and well-defined ligand structures. In order for the benzodiazepines to bind to these sites, an alkyl group of 3 carbons or less is required at position 1, together with a carbonyl group at position 2. In addition, the binding affinity is enhanced by 4'-halogen or 4'-methoxy substituents, and by 7- or 2'-halogens. However, the affinity is decreased by substitutions at position 3 or 4. Central-type benzodiazepine binding in brain, on the other hand, is not affected by 1-alkyl groups and is inhibited by 4'-substitutions. Furthermore, unlike the peripheral-type site, the brain receptor is stereo-selective. These results demonstrate that the two binding sites are fundamentally different in their recognition of benzodiazepine ligands.