摘要
目的采用冷冻干燥技术制备均匀性和稳定性符合要求的阪崎克罗诺杆菌标准物质,用于实验室内部质量控制。方法对使用菌株阪崎克罗诺杆菌(45403)的生化特征、16SrRNA序列、质谱特征进行确认。采用冷冻干燥技术制备含量为103 CFU/样品的菌球。参照CNAS-GL29《标准物质/标准样品定值的一般原则和统计方法》,对20件样品进行均匀性检验,采用单因素方差分析对结果进行统计分析。将样品分别于-20、2、37℃条件下保藏,对其储藏稳定性和运输稳定性进行评价。组织5家实验室进行协同标定。使用20件婴儿配方乳粉作为基质对阪崎克罗诺杆菌标准物质的使用效果进行确认。结果阪崎克罗诺杆菌(45403)生化鉴定结果、16SrRNA序列的NCBIGenebank比对结果、质谱鉴定结果均为阪崎克罗诺杆菌(Cronobacter sakazakii)。采用单因素方差分析进行均匀性检验, F=1.067, P=0.442,符合标准物质的要求。于-20℃保藏170 d,于25、37℃保藏14d后,样品仍然稳定。经5家实验室协同标定,样品含量均为103 CFU/样品。20件婴儿配方乳粉作为基质加入阪崎克罗诺杆菌标准物质进行检验,均可以检出。结论阪崎克罗诺杆菌(CMCC45403)的生化特征、16SrRNA序列分析、蛋白飞行质谱特征均符合克罗诺杆菌属的典型特征。均匀性、储藏稳定性、运输稳定性及适用性的验证实验结果均符合标准物质的要求。
Objective To develop the Cronobactersakazakii standard substance with uniformity and stability by freeze-drying technique,which is used for internal quality control in the laboratory.Methods The biochemical,16 SrRNA sequence and mass spectrometric characteristics of the strain Cronobactersakazakii(45403) were confirmed.Microspheres with a content of 103 CFU/sample was developed by freeze-drying technique.Refering to CNAS-GL29 Reference materials-General and statistical principlesfor certification,uniformity tests were performed on 20 samples,and the results were statistically analyzed by one-way ANOVA.The samples were stored at-20,2,and 37 ℃,respectively,and their storage stability and transport stability were evaluated.Five laboratories were organized for collaborative calibration.Using 20 infant formulasas substrate,the use effect of the standard substance of Cronobactersakazakii was confirmed.Results The biochemical identification of Cronobactersakazakii(45403),the NCBI genebank alignment of 16 SrRNA sequences,and the results of mass spectrometry were all Cronobactersakazakii.The homogeneity was analyzed by one-way ANOVA,F=1.067,P=0.442,and the homogeneity met the requirements of standard materials.The sample was still stable when storing at-20 ℃ for 170 d and storing at 25 ℃,37 ℃ for 7 d.The samples were co-calibrated by 5 laboratories and the sample content was 103 CFU/sample.Twenty infant formulas with Cronobactersakazakiiall could be checked out all.Conclusion The biochemical characteristics,16 SrRNA sequence analysis and protein flight mass spectrometry of Cronobactersakazakii(CMCC45403) developed in this research are consistent with the typical characteristics of Cronobacter.Its uniformity,storage stability,transport stability and suitability are in accordance with the requirements of standard materials.
作者
骆海朋
瞿洪仁
申静云
任秀
陈怡文
谢冠东
余文
刘娜
丁宏
崔生辉
LUO Hai-Peng;QU Hong-Ren;SHEN Jing-Yun;REN Xiu;CHEN Yi-Wen;XIE Guan-Dong;YU Wen;LIU Na;DING Hong;CUI Sheng-Hui(Natiional Institute for Food and Drug Control,Beijing 100050,China)
出处
《食品安全质量检测学报》
CAS
2019年第1期54-59,共6页
Journal of Food Safety and Quality
基金
科技部"食品安全关键技术研发"重点专项项目(2017YFC1601400)~~
作者简介
骆海朋,硕士,副主任检验技师,主要研究方向为食品微生物检验。E-mail:haipengluo123@sina.com;通讯作者:崔生辉,研究员,主要研究方向为食品安全检测。E-mail:cuishenghui@aliyun.com.
