Comparison of different methods for LDL isolation and radioiodination on liver LDL receptor binding in vitro

Int J Rad Appl Instrum B. 1991;18(5):513-7. doi: 10.1016/0883-2897(91)90112-x.

Abstract

Lipoproteins were isolated either by immunoaffinity chromatography (LDL and VLDL) or ultracentrifugation (LDL). Purified lipoproteins were labeled with 123I using either Iodogen or iodine-monochloride (IC1) each followed by purification with gel-chromatography or dialysis (total of 4 combinations). Lipoprotein-concentrations of 0.1-6 micrograms protein/mL were used for direct binding assays investigating the specific binding of labeled lipoproteins (in the presence of a 50-fold excess of unlabeled lipoproteins) to human liver apo-B, E-receptors. In separate experiments displacement of bound 123I-lipoproteins (labeled by the methods mentioned) by unlabeled ones was studied. The binding capacities estimated by Scatchard analysis were similar to each other (141-163 ng protein bound/mg liver plasma membrane protein) independent from the method used for isolation and labeling. Also the affinity constants were very similar and ranged from 0.9 to 1.7 micrograms protein/L. It is concluded that immunoaffinity chromatography or ultracentrifugation for isolation of lipoproteins and the Iodogen or IC1-method for radiolabeling can be recommended to be equally good for in vitro receptor investigation.

Publication types

  • Comparative Study

MeSH terms

  • Adult
  • Apolipoproteins B / metabolism
  • Cell Membrane / metabolism
  • Chromatography, Affinity
  • Chromatography, Gel
  • Female
  • Humans
  • In Vitro Techniques
  • Iodine Radioisotopes
  • Isotope Labeling
  • Kinetics
  • Lipoproteins, LDL / isolation & purification*
  • Lipoproteins, LDL / metabolism
  • Liver / metabolism*
  • Male
  • Membranes / metabolism
  • Receptors, LDL / metabolism*
  • Ultracentrifugation

Substances

  • Apolipoproteins B
  • Iodine Radioisotopes
  • Lipoproteins, LDL
  • Receptors, LDL