Li Xiaojun, Qi Xiulei, Liu Xingguo, Zhu Jun, Hu Lianghai
Center for Supramolecular Chemical Biology, State Key Laboratory of Supramolecular Structure and Materials, School of Life Sciences, Jilin University, Changchun 130012, China.
Jingjie PTM BioLab Co. Ltd., Hangzhou 310018, China.
Anal Chem. 2024 Dec 10;96(49):19803-19811. doi: 10.1021/acs.analchem.4c05288. Epub 2024 Nov 21.
Outer membrane vesicles (OMVs) secreted by bacteria are emerging diagnostic markers for bacterial infection or disease detection due to their carriage of various signaling molecules. However, actual biological samples of patients are extremely complex, and applying OMVs to clinical diagnosis remains a major challenge. One of the major challenges is that there are still great difficulties in the enrichment of OMVs including tedious steps and lower concentration. And some commonly used exosome enrichment methods, such as ultracentrifugation, still have some shortcomings. Herein, we introduce lipopolysaccharide (LPS) molecularly imprinted polymer (MIP) for efficient capturing and analyzing OMVs, enabling a novel approach to bacterial disease diagnosis based on biorecognition materials. LPS, as a unique structure of Gram-negative bacteria, also widely expressed on the surface of OMVs, which will form cyclic hydrogen bonds with functional monomers of MIP with affinity interactions. The prepared MIP efficiently can isolate OMVs from 100 μL of culture broth via specific affinity LPS in less than 40 min with a recovery rate of over 95%. Moreover, MIP exhibits good reusability, with almost identical enrichment performance after 5 repeated cycles, contributing to reducing experimental costs in both time and economy. The captured OMVs can be detected using Western blotting with target protein antibodies or in combination with proteomic analysis, providing a proteomic biomarker platform for early bacteria disease diagnosis.
细菌分泌的外膜囊泡(OMVs)由于携带各种信号分子,正成为细菌感染或疾病检测的新兴诊断标志物。然而,患者的实际生物样本极其复杂,将OMVs应用于临床诊断仍然是一项重大挑战。主要挑战之一是OMVs的富集仍然存在很大困难,包括步骤繁琐和浓度较低。一些常用的外泌体富集方法,如超速离心,仍然存在一些缺点。在此,我们引入脂多糖(LPS)分子印迹聚合物(MIP)用于高效捕获和分析OMVs,从而实现一种基于生物识别材料的细菌疾病诊断新方法。LPS作为革兰氏阴性菌的独特结构,也广泛表达于OMVs表面,它将通过亲和相互作用与MIP的功能单体形成环状氢键。制备的MIP能够在不到40分钟的时间内,通过与LPS的特异性亲和力,从100μL培养液中高效分离OMVs,回收率超过95%。此外,MIP具有良好的可重复使用性,在5次重复循环后富集性能几乎相同,有助于在时间和经济上降低实验成本。捕获的OMVs可以使用针对目标蛋白的抗体通过蛋白质印迹法进行检测,或者与蛋白质组学分析相结合,为早期细菌疾病诊断提供一个蛋白质组学生物标志物平台。