基于金属有机框架负载铂纳米粒子标记免疫方法检测脂多糖

Detection of lipopolysaccharide using metal-organic framework-supported platinum nanoparticle-labeled immunoassay

中文关键词:  脂多糖  外膜囊泡  金属有机框架  纳米酶  酶联免疫吸附法

英文关键词:lipopolysaccharide  outer membrane vesicles  metal-organic framework  nanozyme  enzyme-linked immunosorbent assay

中文摘要:

      目的 基于金属有机框架负载铂纳米粒子标记免疫方法检测脂多糖(lipopolysaccharide, LPS)。方法 通过外膜囊泡(outer membrane vesicles, OMVs)免疫小鼠获得LPS的多克隆抗体, 并制备由金属有机框架负载铂纳米粒子(platinum nanoparticles, Pt NPs)的复合纳米酶作为抗体的信号标记物结合酶联免疫吸附法(enzyme-linked immunosorbent assay, ELISA), 进一步建立针对LPS的新型免疫传感器。结果 免疫获得效价为1:256000的LPS抗体。在构建的基于复合纳米酶的比色免疫分析方法中, 检测LPS的结果显示: 检出限为5 ng/mL, 较使用天然酶标记抗体的ELISA方法灵敏度提高了4倍, 线性检测范围为20~2000 ng/mL, 且具有良好的稳定性和特异性, 在苹果汁和啤酒中回收率为90.38%~105.71%。结论 本研究研制了特异性强、效价高的LPS抗体, 建立的Pt@MIL101-NH2-ELISA方法具有良好的灵敏度和稳定性, 为LPS的免疫检测提供了新思路。

英文摘要:

      Objective To detection of lipopolysaccharide (LPS) by platinum nanoparticle labeling immunoassay based on metal-organic framework. Methods LPS polyclonal antibodies were obtained through immunization of mice with outer membrane vesicles (OMVs). A composite nanozyme consisting of platinum nanoparticles (Pt NPs) loaded on metal-organic framework (MIL101-NH2) was prepared as a signal label. This nanozyme was integrated with enzyme-linked immunosorbent assay (ELISA) to establish a novel immunosensing platform for LPS detection. Results The prepared LPS antibodies demonstrated a titer of 1:256000. The constructed nanozyme-based colorimetric immunoassay showed the limit of detection was 5 ng/mL (4-fold improvement in sensitivity compared with conventional enzyme-labeled ELISA), linear detection range of 20–2000 ng/mL, and satisfactory stability and specificity. Recoveries in apple juice and beer samples ranged from 90.38% to 105.71%. Conclusion This study develop LPS antibodies exhibited high specificity and strong titer. The establish Pt@MIL101-NH2-ELISA method demonstrated enhanced sensitivity and reliability, providing a new approach for immunological detection of LPS.