Development of sandwich ELISA for detection of soluble advanced oxidative protein products
10.3969/j.issn.1000-484X.2025.05.031
- VernacularTitle:检测可溶性晚期氧化蛋白产物的夹心ELISA的建立
- Author:
Xiaorui HOU
1
;
Ping ZHU
;
Yonghui GUO
;
Ning FU
;
Beiyi LIU
Author Information
1. 南方医科大学基础医学院免疫学教研室,广州 510515
- Publication Type:Journal Article
- Keywords:
Advanced oxidation protein products(AOPPs);
Monoclonal antibody;
Oxidative stress;
ELISA
- From:
Chinese Journal of Immunology
2025;41(5):1208-1214
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To develop a double-antibody sandwich ELISA for detecting soluble advanced oxidation protein products(AOPPs).Methods:BALB/c mice were immunized with sodium hypochlorite-oxidized mouse albumin to generate AOPPs-specific monoclonal antibodies(mAbs).Specificity of the mAbs was assessed using indirect ELISA and Western blot.Competitive ELISA was employed to determine if the epitope recognized by newly prepared mAb was consistent with that recognized by mAb 3F2,which developed in our previous work.The sandwich ELISA was then established,and its specificity and sensitivity were compared with the chloramine-T method,the repeatability of double-mAb sandwich ELISA was verified.Results:A mAb AP-4C5 with specific AOPPs recognition was obtained,two sandwich ELISA were developed for the specific detection of soluble AOPPs.Double-mAb sandwich ELISA,using mAb 3F2 as the capture antibody and mAb AP-4C5 as the detection antibody,detected AOPPs in range of 0.25～2 μg/ml(R2=0.991 80).PcAb-mAb sandwich ELISA,using goat anti-HSA polyclonal antibody as the capture antibody and AP-4C5 as detection antibody,detected AOPPs in range of 1.5～25 μg/ml(R2=0.968 75).The double-mAb sandwich ELISA was found to be more sensitive and specific compared to chloramine-T method.Double-mAb sandwich ELISA has good reproducibility(intra-assay CV:3.23％～4.51％,inter-assay CV:3.08％～5.29％).Conclusion:Two kinds of sandwich ELISA for detecting soluble AOPPs have been estab-lished,which hold promise for the detection of clinical samples and understanding of the pathogenic mechanisms of AOPPs.
