Radioimmunoassay and characterization of enkephalins in rat tissues

Journal Article · Wed Jan 25 04:00:00 EST 1978 · J. Biol. Chem.; (United States)

OSTI ID:6934508

Miller, R J; Chang, K J; Cooper, B; Cuatrecasas, P

A highly sensitive and specific radioimmunoassay for three enkephalins (opiate-like pentapeptides) has been developed. The assay utilizes /sup 125/I-labeled enkephalins and antisera raised in rabbits, to synthetic enkephalins coupled with glutaraldehyde to bovine serum albumin. These antisera show <1% cross-reactivity between H/sub 2/N-Tyr-Gly-Gly-Phe-Met-OH ((Met/sup 5/)enkephalin) and H/sub 2/N-Tyr-Gly-Gly-Phe-Leu-OH ((Leu/sup 5/)enkephalin) and even lower cross-reactvity to ..beta..-endorphin. Morphine shows no cross reactivity at all. The assay can detect as little as 10 fmol of enkephalin. Dose response curves for synthetic enkephalin and enkephalin-like immunoreactivity in acid extracts of brain are superimposable. Enkephalin-like immunoreactivity in tissue extracts is destroyed by treating extracts with leucine aminopeptidase or carboxypeptidase, enzymes which also destroy synthetic enkephalin. An opiate receptor binding assay based on the binding of /sup 125/I-labeled (DAla/sup 2/Leu/sup 5/)-enkephalin to N4TG1 neuroblastoma cells has also been developed. Gel filtration of acid extracts of brain or pituitary gland shows that both contain opiate-like material. However, whereas in brain most of opiate-like material co-chromatographs with enkephalin, in the pituitary no opiate-like material was observed to co-chromatograph with enkephalin but was of higher molecular weight. Enkephalin is widely distributed in the brain. High concentrations of both (Met/sup 5/)- and (Leu/sup 5/)enkephalins are found in the striatum (approximately 5 pmol/mg of protein), lower concentrations in the thalamus and midbrain, and very low concentrations in the cerebellum. The ratio of (Met/sup 5/)enkephalin to (Leu/sup 5/)enkephalin also differs in different brain areas. The ratio is very high in the hippocampus (15.2) and hypothalamus (13.6), but lower in other areas such as the cortex (1.4).

• Research Organization: Wellcome Research Labs., Research Triangle Park, NC
• OSTI ID: 6934508
• Journal Information: J. Biol. Chem.; (United States), Journal Name: J. Biol. Chem.; (United States) Vol. 253:2; ISSN JBCHA
• Country of Publication: United States
• Language: English

Related Subjects

550601* -- Medicine-- Unsealed Radionuclides in Diagnostics
551001 -- Physiological Systems-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
62 RADIOLOGY AND NUCLEAR MEDICINE
ANIMALS
BETA DECAY RADIOISOTOPES
BODY
BRAIN
CENTRAL NERVOUS SYSTEM
CHROMATOGRAPHY
DAYS LIVING RADIOISOTOPES
DISTRIBUTION
ELECTRON CAPTURE RADIOISOTOPES
ENDOCRINE GLANDS
ENKEPHALINS
GLANDS
IMMUNE SERUMS
INTERMEDIATE MASS NUCLEI
IODINE 125
IODINE ISOTOPES
ISOTOPE APPLICATIONS
ISOTOPES
LABELLED COMPOUNDS
MAMMALS
NERVOUS SYSTEM
NUCLEI
ODD-EVEN NUCLEI
ORGANIC COMPOUNDS
ORGANS
PEPTIDES
PITUITARY GLAND
POLYPEPTIDES
PROTEINS
RADIOASSAY
RADIOIMMUNOASSAY
RADIOISOTOPES
RATS
RODENTS
SEPARATION PROCESSES
TISSUE DISTRIBUTION
TRACER TECHNIQUES
VERTEBRATES