Faculty of Pharmaceutical Sciences, Kinki University, 3-4-1 Kowakae, Higashi-osaka, Osaka 577-8502, Japan.
Analyst. 2012 May 7;137(9):2211-7. doi: 10.1039/c2an16015c. Epub 2012 Mar 21.
A lectin-impregnated gel was fabricated at the channel crossing point in a microfluidic chip made from polymethyl methacrylate (PMMA). The acrylamide containing lectin was photopolymerized to form a round gel (radius 60 μm) by irradiation with an argon laser, which was also used for fluorometric detection. This gel was applied to specific concentration, elution, and electrophoretic separation of fluorescent-labeled oligosaccharides. Because the lectin in the polyacrylamide gel was mechanically immobilized, it maintained its activity. The lectin was used to trap up to a few tens of femtomoles of specific oligosaccharides labeled with 8-aminopyrene-1,3,6-trisulfonic acid with 2 min by a factor >800, and the amount trapped corresponded to ca. 70% of lectin in the gel. The trapped oligosaccharides were released from the gel by lowering the pH with an acidic background electrolyte. The oligosaccharides that eluted as a broad band were concentrated by transient isotachophoresis stacking using concentrated sodium borate buffer (pH 11.0). The stacked sample components were then separated and fluorometrically detected at the end of the separation channel. Under the optimized conditions, resolution of the saccharides was good, and was similar to that obtained by pinched injection. The method was applied to preconcentration and analysis of oligosaccharides derived from some glycoproteins.
在聚甲基丙烯酸甲酯（PMMA）微流控芯片的通道交叉点处制备了一种凝集素浸渍的凝胶。含有丙烯酰胺的凝集素通过氩激光照射光聚合形成圆形凝胶（半径 60μm），该激光也用于荧光检测。该凝胶用于特定浓度、洗脱和荧光标记寡糖的电泳分离。由于聚丙烯酰胺凝胶中的凝集素被机械固定，因此它保持了其活性。该凝集素可在 2 分钟内通过因子>800 捕获多达数十飞摩尔用 8-氨基芘-1,3,6-三磺酸标记的特定寡糖，并且捕获的寡糖对应于凝胶中约 70%的凝集素。通过用酸性背景电解质降低 pH 值，可以将寡糖从凝胶中释放出来。用浓缩的硼酸钠缓冲液（pH 11.0）进行瞬时等速电泳堆积，将洗脱为宽带的寡糖浓缩。然后在分离通道末端分离和荧光检测堆积的样品成分。在优化的条件下，糖的分辨率良好，与夹流注射获得的分辨率相似。该方法应用于一些糖蛋白衍生的寡糖的预浓缩和分析。
