该研究制备并检测了不含基质的副溶血性弧菌标准物质。用质谱、生化、16S RNA基因序列测定3种方法对菌株CMCC20030分别确认种属。先采用冷冻干燥技术制备800个样品[103 CFU/样品(20 μL)]。然后随机抽取20个样品进行均匀性检验；再将样品存放25、37 ℃分别在1、3、5、7 d取出进行运输稳定性检验，将样品存放4 ℃分别于7、14、28 d进行短期保藏性检验，将样品存放-20 ℃分别于14、28、60 d的保藏稳定性检验。组织5家实验室对样品进行协同标定。最后用食品基质检测使用效果。菌株CMCC20030经3种方法均鉴定为副溶血性弧菌。均匀性检验中，F样品=1.930，小于FINV（0.05，19，20）。稳定性检验中，于-20 ℃保藏60 d、4 ℃保存28 d、25 ℃保藏7 d和37 ℃保藏3 d后，活菌含量103 CFU/样品。协同标定实验中，5家实验室结果均为103 CFU/样品。使用效果实验中，20种食品基质加入样品后均可以检出，本底对照均未检出。制备的不含基质的副溶血性弧菌样品满足标准物质要求，可依据不同目的灵活使用。

In this study, Vibrio parahaemolyticus reference materials without matrix were prepared and detected. The standard strain CMCC20030 was identified by MALDI-TOF MS, biochemical reaction and 16S RNA gene sequencing. The Vibrio parahaemolyticus samples (800) with 103 CFU/sample (20 μL) were prepared by freeze-drying technology. 20 samples were selected randomly for uniformity test. The samples were stored at 25 ℃ and 37 ℃ for 1, 3, 5 and 7 days respectively for transportation stability test, stored at 4 ℃ for 7, 14 and 28 days respectively for short-term preservation stability test, and stored at -20 ℃ for 14, 28 and 60 days respectively for preservation stability test. Five laboratories were organized for collaborative calibration. 20 food products were chosen as the substrate, the applicability was tested according to GB4789.7-2013. The strain CMCC20030 was identified as Vibrio parahaemolyticus by MALDI-TOF MS, biochemical reaction and 16s RNA gene sequencing. The uniformity result was Fsample=1.930< FINV (0.05, 19, 20) by one-way anova. The samples were still 103 CFU/sample when storing 60 d at -20 ℃, 28 d at 4 ℃, 3 d at 37 ℃ and 7 d at 25 ℃. All the results of 5 laboratories were 103 CFU/sample. All 20 samples presented Vibrio parahaemolyticus positively when adding any substrate, only 20 substrates presented negatively. The Vibrio parahaemolyticus sample without matrix meets relevant standards of uniformity, stability, collaborative calibration and applicability, can be applied to different study as reference material according to different purposes.

国家重点研发计划项目（2018YFC1604303）