Premium
Demonstration of a high density lipoprotein (HDL)‐binding protein in Hep G2 cells using colloidal gold‐HDL conjugates
Author(s) -
Kambouris Ambrosios M.,
Roach Paul D.,
Nestel Paul J.
Publication year - 1988
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(88)80666-5
Subject(s) - conjugate , lipoprotein , high density lipoprotein , chemistry , low density lipoprotein receptor related protein 8 , colloid , hep g2 , biochemistry , cholesterol , very low density lipoprotein , in vitro , organic chemistry , mathematical analysis , mathematics
Solubilized membrane proteins of Hep G2 cells were electrophoretically separated on polyacrylamide gels and electrotransferred onto nitrocellulose paper. Overlaying the nitrocellulose with human high density lipoproteins conjugated to colloidal gold revealed the presence of a single protein band with an apparent molecular mass of 80 kDa. Binding of the conjugates to this protein was specific for high density lipoproteins in as much as it was effectively displaced by an excess of unlabelled high density lipoproteins but not by a similar excess of unlabelled low density lipoproteins. Binding was not dependent on Ca 2+ as 10 mM EDTA had no effect. The binding activity of the solubilized membranes was increased by incubating the cells with non‐lipoprotein cholesterol. This was detected on electroblots and quantified with a new dot blot assay using the colloidal gold‐high density lipoprotein conjugates.