GENERATION AND INITIAL CHARACTERIZATION OF A NOVEL POLYCLONAL ANTIBODY-DIRECTED AGAINST HOMOCYSTEINE THIOLACTONE-MODIFIED LOW-DENSITY-LIPOPROTEIN

Citation
E. Ferguson et al., GENERATION AND INITIAL CHARACTERIZATION OF A NOVEL POLYCLONAL ANTIBODY-DIRECTED AGAINST HOMOCYSTEINE THIOLACTONE-MODIFIED LOW-DENSITY-LIPOPROTEIN, Journal of lipid research, 39(4), 1998, pp. 925-933
Citations number
36
Categorie Soggetti
Biology
Journal title
ISSN journal
00222275
Volume
39
Issue
4
Year of publication
1998
Pages
925 - 933
Database
ISI
SICI code
0022-2275(1998)39:4<925:GAICOA>2.0.ZU;2-P
Abstract
Elevated plasma homocysteine (homocysteinemia) are presumed to be resp onsible for the development of coronary artery disease, however, the p recise etiology is unclear. We examined the possibility that the adduc t formed from the reaction between homocysteine thiolactone, a metabol ic product of homocysteine, and apolipoprotein B-100 lysyl residues of low density lipoprotein (LDL) was immunogenic. New Zealand White rabb its were immunized with this adduct at 6-week intervals. Antisera coll ected following the 3rd immunization was assayed for antibody titers u sing solid phase ELISA techniques. Titers (defined as the inverse of t he greatest serum dilution in which there was a significant difference (P < 0.05) between the percentage antibody bound from the antiserum a nd the pre-immune serum) were approximately 10(5). In competition-base d ELISAs, homocysteine thiolactone-treated LDL competed for binding wi th the antiserum, as the 50% inhibitory concentration was approximatel y 10 mu g/ml. Neither homocysteine, homocystine (homocysteine disulfid e), nor Cu2+-oxidized LDL competed for binding. LDL in which lysyl res idues were derivatized by acetylation or methylation were not recogniz ed by the antiserum. Homocysteine thiolactone-treated plasma competed for binding to the antiserum, whereas native plasma did not. All lipop rotein fractions from the homocysteine thiolactone-treated plasma comp eted for binding to the antiserum. We conclude that homocysteine thiol actone-modified LDL is highly immunogenic and specific for homocystein e thiolactone-modified lysines. The potential for using this antibody as a diagnostic tool for measuring plasma homocysteine concentrations and the implications for understanding diseases induced by homocystein emia are discussed.