蛋白质芯片检测牛乳中乳铁蛋白条件的优化研究  

The optimizing study of protein microarray detection conditions for bovine lactoferrin in milk

作　　者：殷继永[1] 马欣欣[1] 孙静[1] 黄建[1] 朴玮[1] 李瑾[1] 陈頔[1] 曹秋野[1] 霍军生[1] 

机构地区：[1]中国疾病预防控制中心营养与健康所,北京100050

出　　处：《中国食品卫生杂志》2016年第6期720-724,共5页Chinese Journal of Food Hygiene

基　　金：中国疾病预防控制中心青年基金(2015A202)

摘　　要：目的优选并确定蛋白质芯片检测牛乳中乳铁蛋白（lactoferrin,Lf）的检测条件。方法利用蛋白质芯片技术,采用双抗夹心的方式,通过点样探针选择试验、点样一致性试验、棋盘滴定试验、浓度倍比试验、生物检测限试验及检测下限试验最终优化出针对目标蛋白的最佳检测条件。结果最终确定以Lf鼠单抗75#为点样探针,预点样46点,46点至90点为点样一致性区段;点样探针浓度为0.5 mg/ml,检测抗体效价为1∶2 000,在0.6-612ng/ml之间剂量-反应关系呈现S型曲线,其线性范围在9.56-306 ng/ml之间;检测下限为1.68 ng/ml,生物检测限为3.59 ng/ml;建立了对Lf具有最佳回归系数（r=0.998）的回归方程与标准曲线。结论此研究优化了利用蛋白质芯片技术检测Lf的试验条件,为建立定量检测Lf的蛋白质芯片平台奠定了基础。Objective To optimize and determine the protein microarray detection condition for bovine lactoferrin（ Lf） in milk. Methods Protein microarray technology and sandwich method was used to optimize the detection condition by probe experiment,homogeneity of the spots of the probes experiment,chessboard titration experiment,concentration dilution experiment,limit of detection and biologic limit of detection experiment. Results The 75#mouse monoclonal antibody was chosen as the printing probe,the pre-printing number was 46 points and the printing stable zone was from 46 th to 90 th point; the concentration of probe is 0. 5 mg / ml,the valence of detection antibody was 1∶ 2 000; the dose-response relationship appeared the S curve from 0. 6 to 612 ng / ml and the linearity range was from 9. 56 to 306 ng / ml,respectively;the limit of detection and biologic limit of detection was 1. 68 and 3. 59 ng / ml,respectively; finally,the regression equation with an optimum determination coefficient（ r = 0. 998） and the standard curve were established. Conclusion This study optimized the protein microarray detection condition for bovine Lf in milk,and these detection conditions were the basis of the further study for establishing the relevant protein microarray plat.

关 键 词：蛋白质芯片 乳铁蛋白 优化 检测 牛乳 检测条件 

分 类 号：R155.5[医药卫生—营养与食品卫生学]