Affinity chromatography of solubilized opioid binding sites using CH-Sepharose modified with a new naltrexone derivative

An affinity gel containing a newly synthesized derivative of naltrexone, beta-naltrexyl-6-ethylenediamine (NED), was used to purify opioid binding sites by 300-450-fold. Binding sites solubilized from brains of toad, rat and cow using digitonin in the presence of 0.5 M NaCl are retained by the gel and can be eluted with naloxone (1-3 microM) in yields of 20-25%. The biospecificity of the interaction of opioid binding sites with modified beads is supported by the following: 1) unmodified beads do not retain the binding sites, 2) binding sites prebound with an opioid are not retained, 3) dilution of NED gels with unmodified Sepharose progressively reduced efficiency of retention and 4) specific receptor ligands elute binding sites retained on the NED gels.