Development of new enzyme-linked immunosorbent assay for oxidized lipoprotein(a) by using purified human oxidized lipoprotein(a) autoantibodies as capture antibody.
BACKGROUND
Oxidized Lp(a) [ox-Lp(a)] has been reported to play more potent role than native Lp(a) in atherosclerosis. Ox-Lp(a), autoantibodies, and Lp(a) immune complexes have all been detected in vivo. Thus, the isolation of its autoantibodies and the investigation of ox-Lp(a) may provide a new means to explore the exact pathogenic role of ox-Lp(a). We isolated and identified human autoantibodies against ox-Lp(a) and developed a new ELISA for ox-Lp(a) by using autoantibodies as capture antibody.


METHODS
Ox-Lp(a) autoantibodies were isolated and identified from healthy subjects by affinity chromatography. 2 "sandwich" ELISAs were developed for measuring ox-Lp(a) level, using autoantibodies against ox-Lp(a) or rabbit antiserum against ox-LDL as the capture antibody and quantitating with monoclonal anti-apo(a) enzyme conjugate, respectively. Ox-Lp(a) levels were studied by both the ELISAs in 100 patients with coronary heart disease (CHD) and 100 control subjects.


RESULTS
The isolated ox-Lp(a) autoantibodies reacted with both apo(a) and apoB epitopes of Ox-Lp(a). Compared to control, plasma ox-Lp(a) levels in patients with CHD were significantly increased (ELISA using human autoantibodies: 24.3+/-33.4 vs. 8.4+/-9.3 microg/ml, P<0.0001; ELISA using antibodies against ox-LDL: 13.0+/-13.8 vs. 7.3+/-9.7 microg/ml, P<0.0001, respectively). Furthermore, a significantly positive relationship between ox-Lp(a) levels detected by 2 ELISAs was also found (R=0.78, P<0.0001).


CONCLUSIONS
We isolated human autoantibodies against ox-Lp(a), which can recognize both apo(a) and apoB epitopes of ox-Lp(a). The developed ELISA for ox-Lp(a) by using human auoantibodies may more accurately reflect the state of Lp(a) oxidation in vivo. Ox-Lp(a) levels increase in patients with CHD.
Wang
Junjun
J
Department of Biochemistry, Jinling Hospital, Clinical School of Medicine College, Nanjing University, PR China.
Zhang
Chunni
C
Gong
Jianbin
J
Zhu
Yanfei
Y
Fu
Li
L
Wang
Xiangdong
X
Li
Ke
K
D001323:Autoantibodies / Q000276:immunology* / Q000302:isolation & purification*; D003327:Coronary Disease / Q000097:blood / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006801:Humans; D017270:Lipoprotein(a) / Q000097:blood* / Q000276:immunology*; D008297:Male; D008875:Middle Aged; D010084:Oxidation-Reduction; D012680:Sensitivity and Specificity
D001323:Autoantibodies; D017270:Lipoprotein(a)