该研究采用高毒力持留基因型单增李斯特菌819-2菌株全菌蛋白免疫SPF级Balb/C小鼠制备抗血清，利用免疫蛋白质组学对菌株胞外蛋白质组进行分析，旨在挖掘筛选单增李斯特菌强免疫原性蛋白作为特异性抗体制备的候选抗原。超声法提取819-2菌株全菌蛋白免疫SPF级Balb/C小鼠制备抗血清，四次免疫后经间接ELISA测定效价达1:512 000。脱氧胆酸钠（DOC）-10%（m/V）TCA沉淀法提取单增李斯特菌819-2菌株胞外蛋白，利用免疫蛋白质组学和LC-MS/MS技术挖掘并鉴定具有强免疫反应的蛋白点，结果表明双向电泳图谱成功获得85个蛋白点，并成功鉴定了P60、InlC、MltG、Enolase和假定蛋白YxeA family protein等5个强免疫原性蛋白，研究结果为基于强免疫原性蛋白制备特异性抗体用于食品及其加工环境中单增李斯特菌富集及快速检测技术研制提供了数据基础。

Antiserum was prepared by immunizing SPF-grade Balb/C mice with the whole-cell protein of the highly virulent Listeria monocytogenes 819-2 strain. The extracellular proteome of the strain was analyzed using the immunoproteomics approach, to identify and obtain strong immunogenic proteins of L. monocytogenes as candidate antigens for specific antibody preparation. The whole-cell protein of the 819-2 strain was extracted using the ultrasonic method. The SPF-grade Balb/C mice were immunized four times using this protein extract to prepare antiserum, and the titer of the antiserum equaled 1:512 000 as measured using indirect ELISA. The extracellular protein of L. monocytogenes 819-2 strain was extracted using TCA precipitation with 10% (m/V) deoxycholate sodium (DOC). The spots for strong immunogenic proteins were identified with the help of immunoproteomics and LC-MS/MS. Two-dimensional electrophoresis results showed 85 protein spots and five strong immunogenic proteins, including P60, InlC, MltG, Enolase, and YxeA family protein, were successfully identified. The results provide a foundation for the preparation of specific antibodies based on highly immunogenic proteins for the enrichment and rapid detection of L. monocytogenes in food and food processing environments.

国家自然科学基金项目（32072326；32222068）；广东省基础与应用基础研究基金项目（2022B1515020068）；广东省科学院创新驱动发展能力建设专项资金项目（2021GDASYL-20210103012）