Quinacrine is a noncompetitive antagonist of the nicotinic acetylcholine receptor (AChR) which displays severalfold fluorescent enhancement upon binding to AChR-rich membranes from Torpedo californica electric organ. It is demonstrated that the fluorescence enhancement comprises two components: specific interaction at a high-affinity binding site on the AChR, and interaction with the lipid bilayer. The interaction with the lipid bilayer can be attenuated by other noncompetitive antagonists, but at concentrations substantially higher than those required for binding to the AChR. It is further shown that quinacrine can inhibit the binding of [3H]phencyclidine and [3H]ethidium in a manner fully consistent with simple competitive inhibition. The data support a model for high-affinity quinacrine binding to the same, single locus of the acetylcholine receptor as phencyclidine and ethidium. This site is likely within the lumen of the ion channel.