skip to main content
OSTI.GOV title logo U.S. Department of Energy
Office of Scientific and Technical Information

Title: A sensitive and rapid method for identification and characterization of low abundance receptors

Abstract

An improved method for detection of low intensity radioligand-receptor complexes resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) is described. (3H)Azidopine-labeled 1,4-dihydropyridine (DHP) receptor from skeletal muscle resolved by SDS-PAGE was transferred to nitrocellulose and cut into strips and individual slices were analyzed for radioincorporation by liquid scintillation counting. (3H)Azidopine-labeled DHP binding subunit migrated as a single entity with a mass of 170 kDa and was confirmed using conventional methods. Results were obtained within 4 h after resolution by SDS-PAGE compared to 3-40 days using conventional methods. In addition, detection of extremely low signals (less than 50 cpm/lane), otherwise overwhelmed by background noise using conventional methods, was possible due to removal of free ligand during electro-transfer to nitrocellulose. This technique offers a rapid sensitive, cost effective alternative to fluorography or other conventional gel slice analysis methods for detecting low intensity radiolabeled complexes resolved by SDS-PAGE.

Authors:
; ;  [1]
  1. (Univ. of Ottawa (Canada))
Publication Date:
OSTI Identifier:
7016215
Resource Type:
Journal Article
Resource Relation:
Journal Name: Analytical Biochemistry; (USA); Journal Volume: 185:1
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; PARASYMPATHOMIMETICS; RECEPTORS; CHEMICAL COMPOSITION; AFFINITY; ELECTROPHORESIS; LIGANDS; MUSCLES; PEPTIDE HYDROLASES; RABBITS; SCINTILLATION COUNTING; TRACER TECHNIQUES; TRITIUM COMPOUNDS; ANIMALS; AUTONOMIC NERVOUS SYSTEM AGENTS; COUNTING TECHNIQUES; DRUGS; ENZYMES; HYDROGEN COMPOUNDS; HYDROLASES; ISOTOPE APPLICATIONS; MAMMALS; MEMBRANE PROTEINS; ORGANIC COMPOUNDS; PROTEINS; VERTEBRATES; 550201* - Biochemistry- Tracer Techniques

Citation Formats

Sunahara, R.K., Murphy, B.J., and Tuana, B.S.. A sensitive and rapid method for identification and characterization of low abundance receptors. United States: N. p., 1990. Web. doi:10.1016/0003-2697(90)90269-F.
Sunahara, R.K., Murphy, B.J., & Tuana, B.S.. A sensitive and rapid method for identification and characterization of low abundance receptors. United States. doi:10.1016/0003-2697(90)90269-F.
Sunahara, R.K., Murphy, B.J., and Tuana, B.S.. Thu . "A sensitive and rapid method for identification and characterization of low abundance receptors". United States. doi:10.1016/0003-2697(90)90269-F.
@article{osti_7016215,
title = {A sensitive and rapid method for identification and characterization of low abundance receptors},
author = {Sunahara, R.K. and Murphy, B.J. and Tuana, B.S.},
abstractNote = {An improved method for detection of low intensity radioligand-receptor complexes resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) is described. (3H)Azidopine-labeled 1,4-dihydropyridine (DHP) receptor from skeletal muscle resolved by SDS-PAGE was transferred to nitrocellulose and cut into strips and individual slices were analyzed for radioincorporation by liquid scintillation counting. (3H)Azidopine-labeled DHP binding subunit migrated as a single entity with a mass of 170 kDa and was confirmed using conventional methods. Results were obtained within 4 h after resolution by SDS-PAGE compared to 3-40 days using conventional methods. In addition, detection of extremely low signals (less than 50 cpm/lane), otherwise overwhelmed by background noise using conventional methods, was possible due to removal of free ligand during electro-transfer to nitrocellulose. This technique offers a rapid sensitive, cost effective alternative to fluorography or other conventional gel slice analysis methods for detecting low intensity radiolabeled complexes resolved by SDS-PAGE.},
doi = {10.1016/0003-2697(90)90269-F},
journal = {Analytical Biochemistry; (USA)},
number = ,
volume = 185:1,
place = {United States},
year = {Thu Feb 15 00:00:00 EST 1990},
month = {Thu Feb 15 00:00:00 EST 1990}
}