Sun Xiuhua, Yang Weichun, Pan Tao, Woolley Adam T
Department of Chemistry and Biochemistry, Brigham Young University, Provo, Utah 84602-5700, USA.
Anal Chem. 2008 Jul 1;80(13):5126-30. doi: 10.1021/ac800322f. Epub 2008 May 15.
Immunoaffinity monolith pretreatment columns have been coupled with capillary electrophoresis separation in poly(methyl methacrylate) (PMMA) microchips. Microdevices were designed with eight reservoirs to enable the electrically controlled transport of selected analytes and solutions to carry out integrated immunoaffinity extraction and electrophoretic separation. The PMMA microdevices were fabricated reproducibly and with high fidelity by solvent imprinting and thermal bonding methods. Monoliths with epoxy groups for antibody immobilization were prepared by direct in situ photopolymerization of glycidyl methacrylate and ethylene glycol dimethacrylate in a porogenic solvent consisting of 70% 1-dodecanol and 30% cyclohexanol. Antifluorescein isothiocyanate was utilized as a model affinity group in the monoliths, and the immobilization process was optimized. A mean elution efficiency of 92% was achieved for the monolith-based extraction of fluorescein isothiocyanate (FITC)-tagged human serum albumin. FITC-tagged proteins were purified from a contaminant protein and then separated electrophoretically using these devices. The developed immunoaffinity column/capillary electrophoresis microdevices show great promise for combining sample pretreatment and separation in biomolecular analysis.
免疫亲和整体柱预处理柱已与聚甲基丙烯酸甲酯（PMMA）微芯片中的毛细管电泳分离相结合。微器件设计有八个储液器，以实现选定分析物和溶液的电控传输，从而进行集成免疫亲和萃取和电泳分离。通过溶剂压印和热键合方法可重复且高保真地制造PMMA微器件。通过甲基丙烯酸缩水甘油酯和乙二醇二甲基丙烯酸酯在由70% 1 - 十二烷醇和30%环己醇组成的致孔溶剂中直接原位光聚合制备具有用于固定抗体的环氧基团的整体柱。抗异硫氰酸荧光素被用作整体柱中的模型亲和基团，并对固定过程进行了优化。基于整体柱萃取异硫氰酸荧光素（FITC）标记的人血清白蛋白的平均洗脱效率达到了92%。FITC标记的蛋白质从污染蛋白中纯化出来，然后使用这些器件进行电泳分离。所开发的免疫亲和柱/毛细管电泳微器件在生物分子分析中结合样品预处理和分离方面显示出巨大的潜力。
