单核细胞增生李斯特氏菌标准物质冻干保存方法

Freeze-drying and preservation method studies on Standard Reference Materials of Listeria monocytogenes

通过研究单核细胞增生李斯特氏菌(Listeria monocytogenes)冷冻干燥保护剂的灭菌方式以及冻干样品的复水方式对样品检测结果的影响,研究设计保护剂种类和含量的正交试验,筛选出最优保护剂灭菌方式和冻干样品复水方式,以及最优的保护剂成分组合。通过研究选择冻干程序和真空压盖,可以保证细菌冻干后保护率在30%以上。此外,针对不同菌体密度的样品储存环境进行考察,通过比较冻干菌粉的保护率发现高密度菌体在-20℃储存30d稳定。研究制备的活菌在冷链运输(加冰袋)条件下一周内稳定,满足标准物质的发放运输要求,满足实验室质控和方法验证的基本需求。文章所述的单核细胞增生李斯特菌标准物质研究结果将为其他微生物标准物质的冻干保护方法和稳定性研究提供理论指导。

We study on the sterilization methods of freeze-drying protectant for Listeria monocytogenes and rehydration methods of freeze-dried samples, as well as the orthogonal test of protective agent of differenet types or quantities. As a result, we successfully selected the optimal protective agent sterilization method and the lyophilized sample rehydration method , as well as the combination of protective agent components. The protection rate of bacteria after lyophilization can be guaranteed to be above 30%, through selection of freeze-drying procedures and vacuum capped. In addition, it was found that the highdensity cells were stable at -20 ℃ for one month by comparing the protection rate of the freeze-dried powder. The live bacteria quantity is stable within one week under the condition of cold chain transportation, which will meet the requirements for the issuance and transportation of standard materials, and the basic requirements of laboratory quality control and method validation. The results of the reference material of Listeria monocytogenes in this paper will provide theoretical guidance for the freeze-drying protection methods and stability studies of other microbial reference materials.