Dot-blot assay for the low density lipoprotein receptor
We describe a new method for detecting the interaction of low density lipoprotein with its receptor using unmodified nitrocellulose as support for membrane protein. The method is specific and sensitive down to 3 micrograms of membrane protein. Unlabeled LDL, but not HDL, competes with /sup 125/I-labeled LDL for binding, and binding is abolished by pretreatment of the membranes with pronase and is dependent upon the presence of Ca2+. Furthermore, modification of arginine or lysine residues on LDL abolishes the lipoprotein interaction with the receptor protein supported on the nitrocellulose. When the membranes are solubilized with octyl glucoside, purification steps of the receptor can be directly followed with no interference of the detergent, therefore eliminating the need for its removal. The increased expression of LDL receptors on liver membranes from estradiol-treated rats was also demonstrated. We suggest, therefore, that this method can be used to detect the presence of LDL receptors on minute amounts of membrane protein.
- Research Organization:
- Institute of Pharmacological Sciences, Milano, Italy
- OSTI ID:
- 6528356
- Journal Information:
- J. Lipid Res.; (United States), Journal Name: J. Lipid Res.; (United States) Vol. 1; ISSN JLPRA
- Country of Publication:
- United States
- Language:
- English
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59 BASIC BIOLOGICAL SCIENCES
ALKALINE EARTH METAL COMPOUNDS
ANIMALS
BETA DECAY RADIOISOTOPES
BODY
CALCIUM COMPOUNDS
CATTLE
CPB
DAYS LIVING RADIOISOTOPES
DIGESTIVE SYSTEM
DOMESTIC ANIMALS
ELECTRON CAPTURE RADIOISOTOPES
GLANDS
INTERMEDIATE MASS NUCLEI
IODINE 125
IODINE ISOTOPES
ISOTOPE APPLICATIONS
ISOTOPE DILUTION
ISOTOPES
LIPIDS
LIPOPROTEINS
LIVER
MAMMALS
MEASURING METHODS
MEMBRANE PROTEINS
NUCLEI
ODD-EVEN NUCLEI
ORGANIC COMPOUNDS
ORGANS
PROTEINS
RADIOISOTOPES
RATS
RECEPTORS
RODENTS
RUMINANTS
SPECIFICITY
TRACER TECHNIQUES
VERTEBRATES