Irish Separation Science Cluster, National Centre for Sensor Research and School of Chemical Sciences, Dublin City University, Dublin, Ireland.
Electrophoresis. 2010 Nov;31(22):3783-6. doi: 10.1002/elps.201000457.
Fluorescently labeled carbohydrates released from glycoproteins were separated using a commercially available microfluidic chip electrophoresis system. While the instrumentation was primarily designed for DNA analysis it was found that the application base can be easily expanded using the development software provided by the manufacturer. The carbohydrates were released by enzymatic digestion (PNGase F) from glycoproteins present in human plasma after boronic acid - lectin affinity enrichment. After fluorescent labeling with 8-aminopyrene-1,3,6-trisulfonic acid the carbohydrates were separated based on capillary gel electrophoresis mechanism and detected by a fluorescence detector using a blue (470 nm) LED. The separation was completed in 40 s in a microfluidic channel of 14 mm length. Glucose ladder carbohydrate oligomers differing by one glucose unit were baseline separated up to a 20-mer with the main limitation being the detection sensitivity. As expected, the observed resolution in these experiments did not approach that of standard CE with 20 times longer separation distance; however, the chip-based analysis excelled in the speed of the separation. Similar electrophoretic profiles of glycans released from plasma glycoproteins were obtained using a standard CE equipment with 35 cm separation length and microfluidic chips with a separation distance of only 14 mm.
从糖蛋白中释放的荧光标记碳水化合物使用市售的微流控芯片电泳系统进行分离。虽然该仪器主要设计用于 DNA 分析，但发现使用制造商提供的开发软件可以轻松扩展其应用基础。碳水化合物通过酶消化（PNGase F）从硼酸 - 凝集素亲和富集的人血浆中的糖蛋白中释放。用 8-氨基芘-1,3,6-三磺酸进行荧光标记后，根据毛细管凝胶电泳机制进行分离，并使用蓝色（470nm）LED 的荧光检测器进行检测。在 14mm 长的微流道中，40s 即可完成分离。葡萄糖阶梯碳水化合物低聚糖相差一个葡萄糖单位，基线分离到 20 个低聚糖，主要限制是检测灵敏度。正如预期的那样，在这些实验中观察到的分辨率并没有接近分离距离长 20 倍的标准 CE 的分辨率；然而，基于芯片的分析在分离速度方面表现出色。使用标准 CE 设备（分离长度为 35cm）和分离距离仅为 14mm 的微流控芯片，可获得从血浆糖蛋白中释放的聚糖的相似电泳图谱。
