Simultaneous detection of α-Lactoalbumin, β-Lactoglobulin and Lactoferrin in milk by Visualized Microarray

Zhoumin Li, Fang Wen, Zhonghui Li, Nan Zheng, Jindou Jiang, Danke Xu

α-Lactalbumin (a-LA), β-lactoglobulin (β-LG) and lactoferrin (LF) are of high nutritional value which have made ingredients of choice in the formulation of modern foods and beverages. There remains an urgent need to develop novel biosensing methods for quantification featuring reduced cost, improved sensitivity, selectivity and more rapid response, especially for simultaneous detection of multiple whey proteins. A novel visualized microarray method was developed for the determination of a-LA, β-LG and LF in milk samples without the need for complex or time-consuming pre-treatment steps. The measurement principle was based on the competitive immunological reaction and silver enhancement technique. In this case, a visible array dots as the detectable signals were further amplified and developed by the silver enhancement reagents. The microarray could be assayed by the microarray scanner. The detection limits (S/N = 3) were estimated to be 40 ng/mL (α-LA), 50 ng/mL (β-LG), 30 ng/mL (LF) (n = 6). The method could be used to simultaneously analyze the whey protein contents of various raw milk samples and ultra-high temperature treated (UHT) milk samples including skimmed milk and high calcium milk. The analytical results were in good agreement with that of the high performance liquid chromatography. The presented visualized microarray has showed its advantages such as high-throughput, specificity, sensitivity and cost-effective for analysis of various milk samples.