Preparation and application of monoclonal antibodies against oxidized DJ-1. Significant elevation of oxidized DJ-1 in erythrocytes of early-stage Parkinson disease patients.
DJ-1 was initially identified as a novel oncogene and has recently been found to be a causative gene for a familial form of Parkinson's disease (PD), viz, PARK7. Cysteine residue at position 106 (Cys-106) in DJ-1 was found to be oxidized preferentially under oxidative stress. In the present study, we developed specific antibodies against Cys-106-oxidized DJ-1 using baculovirus particles displaying the surface glycoprotein gp64-fusion protein as the immunizing agent. Western blot analysis combined with two-dimensional gel electrophoresis revealed that these antibodies specifically recognized oxidized DJ-1. Furthermore, we developed a competitive enzyme-linked immunosorbent assay (ELISA) for detecting oxidized DJ-1 and measured blood levels of oxidized DJ-1 in PD patients (n=15). It was observed that the levels of oxidized DJ-1 in erythrocytes of unmedicated PD patients were markedly higher without overlap than those of medicated PD patients and healthy subjects. No significant difference was observed in DJ-1 levels between mediated and unmediated PD patient. These results suggest the oxidative modification of DJ-1 in PD patients and the potential application of the antibody for diagnosis of PD at early-stage.
Saito
Yoshiro
Y
Health Technology Research Center, National Institute of Advanced Industrial Science and Technology, Ikeda, Osaka, Japan. ysaito@mail.doshisha.ac.jp
Hamakubo
Takao
T
Yoshida
Yasukazu
Y
Ogawa
Yoko
Y
Hara
Yasuo
Y
Fujimura
Harutoshi
H
Imai
Yasuharu
Y
Iwanari
Hiroko
H
Mochizuki
Yasuhiro
Y
Shichiri
Mototada
M
Nishio
Keiko
K
Kinumi
Tomoya
T
Noguchi
Noriko
N
Kodama
Tatsuhiko
T
Niki
Etsuo
E
D000368:Aged; D000704:Analysis of Variance; D000906:Antibodies / Q000378:metabolism*; D000978:Antiparkinson Agents / Q000494:pharmacology; D016367:Baculoviridae; D015153:Blotting, Western; D045744:Cell Line, Tumor; D015180:Electrophoresis, Gel, Two-Dimensional; D004797:Enzyme-Linked Immunosorbent Assay; D004912:Erythrocytes / Q000378:metabolism*; D005260:Female; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism; D047908:Intracellular Signaling Peptides and Proteins / Q000097:blood / Q000276:immunology* / Q000378:metabolism*; D008297:Male; D008875:Middle Aged; D015513:Oncogene Proteins / Q000097:blood / Q000276:immunology* / Q000378:metabolism*; D010084:Oxidation-Reduction; D010300:Parkinson Disease / Q000097:blood / Q000188:drug therapy / Q000378:metabolism*; D000071617:Protein Deglycase DJ-1; D013997:Time Factors
D000906:Antibodies; D000978:Antiparkinson Agents; D047908:Intracellular Signaling Peptides and Proteins; D015513:Oncogene Proteins; D006861:Hydrogen Peroxide; C105131:PARK7 protein, human; D000071617:Protein Deglycase DJ-1