Skip to main content
PMC home page
The Journal of Clinical Investigation logoLink to The Journal of Clinical Investigation
. 1969 Aug;48(8):1387–1396. doi: 10.1172/JCI106104

Radioimmunoassay of beta lipoprotein—protein of rat serum

R Philip Eaton 1, David M Kipnis 1
PMCID: PMC322365  PMID: 4978731

Abstract

A double antibody radioimmunoassay for rat serum beta lipoprotein-protein (beta Lp-protein) is described. The protein was purified by ultracentrifugation, selective heparin-manganous precipitation, and gel filtration on Sephadex G-200. Antiserum was prepared in rabbits by biweekly immunization and absorbed with nonbeta lipoprotein containing rat serum. Iodination with 125I and purification by gel filtration provided a radiolabeled protein which was > 98% displaced by purified beta lipoprotein in the immunoassay. The radioimmunoassay was sensitive to beta Lp-protein concentrations from 0.1 to 1.5 μg. Specificity of the immunoassay for beta Lp-protein was established by comparison of the displacement curves obtained with serum very low density lipoprotein (VLDL), low density lipoprotein (LDL), high density lipoprotein (HDL), and density (d) > 1.21 fractions and with the beta and alpha migrating lipoproteins eluted from paper electrophoretograms. Suitability of the assay for measuring beta Lp-protein in serum was established by demonstrating 100% recovery of beta lipoprotein added to whole serum and by the absence of immunoreactive beta Lp-protein in serum of orotic acid-treated rats. Examination of sera from six other vertebrates species revealed partial cross-reactivity. Normal rat serum was found to contain 0.25±0.01 mg/ml of beta Lp-protein and hepatic production by an isolated perfused rat liver system was determined as 0.145 mg/hr.

Full text

PDF
1387

Images in this article

1388Image
on p.1388
1389Image
on p.1389

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Aladjem F. Immunoelectrophoretic properties of low-density human serum lipoproteins. Nature. 1966 Mar 5;209(5027):1003–1005. doi: 10.1038/2091003a0. [DOI] [PubMed] [Google Scholar]
  2. Camejo G. Structural studies of rat plasma lipoproteins. Biochemistry. 1967 Oct;6(10):3228–3241. doi: 10.1021/bi00862a032. [DOI] [PubMed] [Google Scholar]
  3. FIREMAN P., VANNIER W. E., GOODMAN H. C. IMMUNOCHEMICAL STUDIES OF HUMAN SERUM FRACTIONATED BY GEL FILTRATION WITH SEPHADEX G-200. Proc Soc Exp Biol Med. 1964 Apr;115:845–849. doi: 10.3181/00379727-115-29054. [DOI] [PubMed] [Google Scholar]
  4. Franzini C. Gel filtration behaviour of human serum lipoproteins. Clin Chim Acta. 1966 Nov;14(5):576–578. doi: 10.1016/0009-8981(66)90179-3. [DOI] [PubMed] [Google Scholar]
  5. GREENWOOD F. C., HUNTER W. M., GLOVER J. S. THE PREPARATION OF I-131-LABELLED HUMAN GROWTH HORMONE OF HIGH SPECIFIC RADIOACTIVITY. Biochem J. 1963 Oct;89:114–123. doi: 10.1042/bj0890114. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. GUSTAFSON A., ALAUPOVIC P., FURMAN R. H. STUDIES OF THE COMPOSITION AND STRUCTURE OF SERUM LIPOPROTEINS: ISOLATION, PURIFICATION, AND CHARACTERIZATION OF VERY LOW DENSITY LIPOPROTEINS OF HUMAN SERUM. Biochemistry. 1965 Mar;4:596–605. doi: 10.1021/bi00879a033. [DOI] [PubMed] [Google Scholar]
  7. Gustafson A., Alaupovic P., Furman R. H. Studies of the composition and structure of serum lipoproteins. Separation and characterization of phospholipid-protein residues obtained by partial delipidization of very low density lipoproteins of human serum. Biochemistry. 1966 Feb;5(2):632–640. doi: 10.1021/bi00866a033. [DOI] [PubMed] [Google Scholar]
  8. Gustafson A. Ultrasoundcardiography in mitral stenosis with particular reference to the relationship to hemodynamic and surgical findings. Acta Med Scand Suppl. 1966;461:1–137. [PubMed] [Google Scholar]
  9. HAFT D. E., ROHEIM P. S., WHITE A., EDER H. A. Plasma lipoprotein metabolism in perfused rat livers. I. Protein synthesis and entry into the plasma. J Clin Invest. 1962 Apr;41:842–849. doi: 10.1172/JCI104541. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. HAVEL R. J., EDER H. A., BRAGDON J. H. The distribution and chemical composition of ultracentrifugally separated lipoproteins in human serum. J Clin Invest. 1955 Sep;34(9):1345–1353. doi: 10.1172/JCI103182. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Katz J., Bonorris G., Okuyama S., Sellers A. L. Albumin synthesis in perfused liver of normal and nephrotic rats. Am J Physiol. 1967 Jun;212(6):1255–1260. doi: 10.1152/ajplegacy.1967.212.6.1255. [DOI] [PubMed] [Google Scholar]
  12. LEES R. S., HATCH F. T. Sharper separation of lipoprotein species by paper electrophoresis in albumin-containing buffer. J Lab Clin Med. 1963 Mar;61:518–528. [PubMed] [Google Scholar]
  13. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  14. MARSH J. B., DRABKIN D. L. Experimental reconstruction of metabolic pattern of lipid nephrosis: key role of hepatic protein synthesis in hyperlipemia. Metabolism. 1960 Oct;9:946–955. [PubMed] [Google Scholar]
  15. MARSH J. B., WHEREAT A. F. The synthesis of plasma lipoprotein by rat liver. J Biol Chem. 1959 Dec;234:3196–3200. [PubMed] [Google Scholar]
  16. MARSH J. B., WHEREAT A. F. The synthesis of plasma lipoprotein by rat liver. J Biol Chem. 1959 Dec;234:3196–3200. [PubMed] [Google Scholar]
  17. RADDING C. M., STEINBERG D. Studies on the synthesis and secretion of serum lipoproteins by rat liver slices. J Clin Invest. 1960 Oct;39:1560–1569. doi: 10.1172/JCI104177. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. STANDERFER S. B., HANDLER P. Fatty liver induced by orotic acid feeding. Proc Soc Exp Biol Med. 1955 Oct;90(1):270–271. doi: 10.3181/00379727-90-22005. [DOI] [PubMed] [Google Scholar]
  19. Schlesinger M. J. The reversible dissociation of the alkaline phosphatase of Escherichia coli. 3. Properties of antibodies directed against the subunit. J Biol Chem. 1967 Apr 10;242(7):1599–1603. [PubMed] [Google Scholar]
  20. WEIGLE W. O. IMMUNOCHEMICAL PROPERTIES OF HEMOCYANIN. Immunochemistry. 1964 Dec;1:295–302. doi: 10.1016/0019-2791(64)90030-8. [DOI] [PubMed] [Google Scholar]
  21. Williamson J. R., Garcia A., Renold A. E., Cahill G. F., Jr Studies on the perfused rat liver. I. Effects of glucagon and insulin on glucose metabolism. Diabetes. 1966 Mar;15(3):183–187. doi: 10.2337/diab.15.3.183. [DOI] [PubMed] [Google Scholar]
  22. Windmueller H. G., Levy R. I. Total inhibition of hepatic beta-lipoprotein production in the rat by orotic acid. J Biol Chem. 1967 May 10;242(9):2246–2254. [PubMed] [Google Scholar]
  23. Zucker L. M. Hereditary obesity in the rat associated with hyperlipemia. Ann N Y Acad Sci. 1965 Oct 8;131(1):447–458. doi: 10.1111/j.1749-6632.1965.tb34810.x. [DOI] [PubMed] [Google Scholar]
  24. ungenberg de Jong J. J., Marsh J. B. Biosynthesis of plasma lipoproteins by rat liver ribosomes. J Biol Chem. 1968 Jan 10;243(1):192–199. [PubMed] [Google Scholar]

Articles from Journal of Clinical Investigation are provided here courtesy of American Society for Clinical Investigation

RESOURCES