Simple and practical sandwich-type enzyme immunoassay for human oxidatively modified low density lipoprotein using antioxidized phosphatidylcholine monoclonal antibody and antihuman apolipoprotein-B antibody.
OBJECTIVE
To develop a simple and practical enzyme immunoassay (EIA) for oxidatively modified low density lipoprotein (Ox-LDL) in human blood, a biological marker of atherogenesis.


METHODS
A sandwich EIA suitable for the measurement of human Ox-LDL was developed using the mouse monoclonal antibody FOH1a/DLH3. This antibody, specific for oxidized phosphatidylcholine, was used as the capture antibody, and a horseradish peroxidase (HRP)-labeled goat anti-human apolipoprotein-B (Apo-B) IgG was used for detection. Copper-oxidized human LDL, prepared under controlled conditions, was used as a standard and the results of the EIA were expressed in arbitrary units (U/mL).


RESULTS
This EIA meets all the requirements for use in routine clinical assays in terms of sensitivity (detection limit: 1 U/mL), reproducibility (total CV: 2.3-7.7%), accuracy (recovery: 90.6-103.8%), simplicity and rapidity (<4 h). Clinical performance of the assay was assessed by measurement of the Ox-LDL in the plasma of normal subjects (10.8 +/- 2.8 U/mL, mean +/- SD) and patients with coronary heart disease (CHD) (19.7 +/- 10.2). The present EIA had a sensitivity of 79% and a specificity of 75% for CHD.


CONCLUSIONS
We have developed a new assay, suitable for the measurement of Ox-LDL in human blood, which meets the requirements for routine clinical assay.
Kohno
H
H
Fuji Research Laboratories, Kyowa Medex Co., Ltd., Nagaizumi-cho, Shizuoka, Japan. hiroaki.kohno@kyowa.co.jp
Sueshige
N
N
Oguri
K
K
Izumidate
H
H
Masunari
T
T
Kawamura
M
M
Itabe
H
H
Takano
T
T
Hasegawa
A
A
Nagai
R
R
D000328:Adult; D000911:Antibodies, Monoclonal / Q000096:biosynthesis; D001055:Apolipoproteins B / Q000097:blood* / Q000276:immunology; D015415:Biomarkers / Q000097:blood; D002850:Chromatography, Gel; D002940:Circadian Rhythm; D003327:Coronary Disease / Q000097:blood / Q000175:diagnosis; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007124:Immunoenzyme Techniques; D008077:Lipoproteins, LDL / Q000097:blood*; D008297:Male; D016555:Matched-Pair Analysis; D008875:Middle Aged; D010713:Phosphatidylcholines / Q000097:blood* / Q000276:immunology; D012372:ROC Curve; D012016:Reference Values
D000911:Antibodies, Monoclonal; D001055:Apolipoproteins B; D015415:Biomarkers; D008077:Lipoproteins, LDL; D010713:Phosphatidylcholines; C099252:oxidized low density lipoprotein