Demonstration of a high density lipoprotein (HDL)‐binding protein in Hep G2 cells using colloidal gold‐HDL conjugates

FEBS Letters

Roach, Paul D.¹  Nestel, Paul J.¹  Kambouris, Ambrosios M.¹ 

[1] CSIRO, Division of Human Nutrition, Kintore Avenue, Adelaide, SA 5000, Australia

关键词: HDL receptor;    Colloidal gold;    Electroblotting;    Dot blotting;    Ligand blotting;    (Hep G2 cell);    HDL;    high density lipoproteins;    LDL;    low density lipoproteins;    VLDL;    very low density lipoproteins;    MEM;    minimal essential medium;    PBS;    phosphate buffered saline;    PMSF;    phenylmethylsulfonyl fluoride;    BSA;    bovine serum albumin;   

DOI  :  10.1016/0014-5793(88)80666-5

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

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 Ca²⁺ 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.

【 授权许可】

Unknown