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Sera containing antibodies to beta-endorphin from 2 patients with major depressive disorder were shown to have antidiotypic antibodies that specifically inhibited reactivity between anti-beta-endorphin IgG and beta-endorphin. Autologous and homologous antiidiotypic anti-anti-beta-endorphin IgG antibodies were isolated by affinity chromatography. The purified antiidiotypic antibody did not bind beta-endorphin but competed with [125I]beta-endorphin for rat brain opiate receptors. Normal IgG that was similarly treated had negligible competitive effects. The antibody bound to the membrane preparation; such binding was inhibited by opiate receptor ligands. Binding of the antiidiotype to a 60,000-dalton protein from rat brain was detected by Western immunoblot analysis. This protein corresponds in molecular weight to proteins proposed to be components of opiate receptors. These findings imply that immune reactivity to neuropeptides could contribute to psychiatric impairment.

Human IgG specific for (8-endorphin was identified by enzyme-linked immunoabsorbent assay and isolated by affinity chromatography. From a sample of 27 subjects, three individuals with major depression demonstrated plasma IgG highly reactive with human 13-endorphin, while four other subjects (two with depression and two randomly selected blood donors) had intermediate reactivity. An enzyme-linked immunoabsorbent assay (ELISA) system was used to detect human and rabbit antibodies to 3-endorphin. In preliminary studies, the sensitivity and specificity of this assay was examined using rabbit anti-pendorphin antiserum (Immuno Nuclear, Stillwater, MN). This antiserum was found to have relatively high specificity for ,-endorphin with negligible reactivity to bovine plasma albumin (<3.5%) or other human neuropeptides (2.5-7.9%). In similar experiments, comparable specificity was found using antisera to other neuropeptides including corticotropin, a-melanotropin, and substance P, firmly establishing that each of these neuropeptides bound to ELISA plates (data not shown). In these preliminary studies, no data were available for human antibodies directed against neuropeptides. Using the rabbit anti-/3-endorphin antibody, we detected lowest reactivity against substance P and corticotropin and decided to use these as background for the human studies. MATERIALS AND METHODSImmunoglobulins. We examined purified IgG after either ammonium sulfate precipitation or protein A (Staphylococcus aureus) column separation. Serum samples were obtained from a general pool of6 blood donors, 9 healthy normal volunteers, and 12 psychiatric patients meeting Diagnostic and Statistical Manual of Mental Disorders III (13) criteria for either major depression (n = 9) or personality disorder (n = 3) who were sequentially admitted over a several-month period to a National Institute of Mental Health Clinical Center psychiatric unit. One of the subjects with major depression (subject 10) had a 1-year history of chronic pain and a prior history of an urticarial reaction to hydromorphone hydrochloride.ELISA. P-Endorphin (10 gg/ml) and substance P or corticotropin (10 ,g/ml) in pH 9.4 coating buffer were pipetted in a 100-,ul volume into an Immulon I polystyrene plate (Dynatech, New York) and allowed to stand at 0C for 48 hr. The plate was washed with 0.5% Tween 20 (Fisher) in phosphate-buffered saline (PBS; pH 7.3, without calcium or magnesium), and nonspecific binding sites were blocked with 100 ,ul of 0.5% bovine plasma albumin in distilled water for 20 min at room temperature. Then 100 41. of human IgG was added and incubated at 370C for 2 hr. The plate was then washed, and 100 ,1u of a 1:200 dilution of an alkaline phosphatase-conjugated goat anti-human Fc IgG (heavychain specific) antibody (Sigma) was pipetted into each well.After 2 hr at 37°C, the plate was washed, 100 ,u1 of the alkaline phosphatase substrate p-nitrophenyl phosphate (disodium salt) dissolved in 10% diethanolamine was added to each well, and the reaction was allowed to pro...

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