Ma Xinxin, Yin Jiyong, Sun Jing, Huang Jian, Piao Wei, Li Jin, Chen Di, Yuan Xiaolin, Huo Junsheng
National Institute for Nutrition and Health, Chinese Center for Diease Control and Prevention, Beijing 100050, China.
Wei Sheng Yan Jiu. 2017 Jan;46(1):78-83.
To optimize the conditions of protein chip assay for bovine milk β-Lactoglobulin( β-Lg).
A microarrayer was used for printing anti-β-Lg as antibody I on each 3-dimensional-slide, another antis β-Lg antibody was used as detection antibody II and goat antibody coupled to Cy3 was used as antibody III. The standard β-Lg was detected by double antibody sandwich technique.
Mouse monoclonal β-Lg antibody66# was chosen as the probe and contact printing as the printing method. The range between 42 and 92 spots was chosen as the basic printing condition. The concentration of β-Lg probes was 0. 5 mg / mL. The β-Lg detection antibody titre was 1∶2000. One percent no protein blocking solution was choosen as the blocking buffer. The lower detection limit and the biological detection limit of β-Lg were 17. 54 ng / m L and 55. 31 ng / m L respectively. The linear range was determined according to the S type curve of β-Lg and the best fitting models and standard curve were established for β-Lg( R~2=0. 9993).
The study optimizes conditions of a quantitative analysis system for measurement of β-Lg with protein chip, thus establishing the protein chip platform for quantitative detection of bovine milk β-Lactoglobulin.
优化牛乳β-乳球蛋白（β-Lg）蛋白芯片检测条件。
使用微阵列点样仪将抗β-Lg作为抗体I点印在每张三维玻片上，另一种抗β-Lg抗体作为检测抗体II，与Cy3偶联的山羊抗体作为抗体III。采用双抗体夹心技术检测标准β-Lg。
选择小鼠单克隆β-Lg抗体66#作为探针，采用接触式点样法。选择42至92个点的范围作为基本点样条件。β-Lg探针浓度为0.5mg/mL。β-Lg检测抗体效价为1∶2000。选择1%无蛋白封闭液作为封闭缓冲液。β-Lg的最低检测限和生物检测限分别为17.54ng/mL和55.31ng/mL。根据β-Lg的S型曲线确定线性范围，并建立了β-Lg的最佳拟合模型和标准曲线（R²=0.9993）。
本研究优化了蛋白芯片定量检测β-Lg的分析系统条件，从而建立了牛乳β-乳球蛋白定量检测的蛋白芯片平台。
