免疫富集联合MALDI-TOF MS检测奶粉中阪崎克罗诺杆菌的方法建立

目的:制备高效且特异性好的阪崎克罗诺杆菌抗体及其免疫磁珠，建立免疫富集联合基质辅助激光解吸电离飞行时间质谱(matrix-assisted laser desorption/ionization time of flight mass spectrometry, MALDI-TOF MS)检测奶粉中的阪崎克罗诺杆菌的方法。方法:制备阪崎克罗诺杆菌混菌抗体及其免疫磁珠，对免疫磁珠分别在纯培养及奶粉基质中的捕获率进行研究，利用MALDI-TOF MS对不同奶粉基质中不同杂菌污染条件下的检测样本进行鉴定并验证免疫磁珠的特异性。结果:阪崎克罗诺杆菌免疫磁珠在纯培养条件及奶粉基质中对4株阪崎克罗诺杆菌及其混菌的捕获率均>80%，联合MALDI-TOF MS鉴定结果显示在奶粉基质中不同杂菌污染条件下具有较好的鉴定结果，即使在高比例（1:100）杂菌污染条件下，仍能准确鉴定奶粉中阪崎克罗诺杆菌，此时检测样品中阪崎克罗诺杆菌浓度仅为20 CFU/mL。结论:本研究建立了一种操作简便、鉴定结果准确的免疫富集联合MALDI-TOF MS检测奶粉中阪崎克罗诺杆菌的方法，为奶粉中阪崎克罗诺杆菌的快速准确鉴定提供了新的方法参考。

Establishment of the Method of Immunoenrichment Combined with MALDI-TOF MS to Detect Cronobacter sakazakii in Milk Powder

Objective:To prepare an efficient and specific antibody against Cronobacter sakazakii and its immunomagnetic beads,and establish a method of immunoenrichment combined with matrix-assisted laser desorption/ionization mass spectrometry(MALDI-TOF MS) for the detection of Cronobacter sakazakii in milk powder.Methods:The antibody and its immune beads of Cronobacter sakazakii were prepared,the capture rate of immune magnetic beads was studied in pure culture and milk powder matrix.The specific of the beads were identified and verified by using MALDI-TOF MS.Results:The capture rate of four strains of Cronobacter sakazakii and its mixed bacteria in the pure culture and milk powder matrix were all over 80%,and the results of MALDI-TOF MS showed that the identification were better under different contamination conditions of different impurities in the milk powder matrix,even under the high proportion of(1:100) of the bacteria,it could be used to identify Cronobacter sakazakii in milk powder accurately,at this time,the concentration of Cronobacter sakazakii in the sample was only 20 CFU/mL.Conclusion:A simple and accurate method for the detection of Cronobacter sakazakii in milk powder by immunoenrichment and MALDI-TOF MS was established,which provided a new method reference for the rapid and accurate identification of Cronobacter sakazakii in milk powder.