Microscale lsoelectric Fractionation Using Photopolymerized Membranes

作者：Sommer Greg J^*; Mai Junyu; Singh Anup K; Hatch Anson V

来源：Analytical Chemistry, 2011, 83(8): 3120-3125.

DOI：10.1021/ac200073p

In this work, we introduce microscale isoelectric fractionation (mu IF) for isolation and enrichment of molecular species at any desired location in a microfluidic chip. Narrow pH specific polyacrylamide membranes are photopatterned in situ for customizable d(evice fabrication; multiple membranes of precise pH are easily incorporated throughout existing channel layouts. Samples are electrophoretically driven across the membranes such that charged species, for example, proteins and peptides, are rapidly discretized, into fractions based on their isoelectric points (pI) without the use of carrier aimpholytes. This format makes fractions easy to compartmentalize and access for integrated preparative or analytical operations on chip We present and discuss the key design considerations and trade-offs associated with proper system operation and optimal run conditions. Efficient and reproducible fractionation of model fluorescent pI markers and proteins is achieved using single membrane fractionators at pH 6.5 and 5.3 from both buffer and Escherichia coli cell lysate sample conditions. Effective fractionation is also shown using a serial 3-membrane fractionator tailored for isolating analytes-of-interest from high abundance components of serum. We further demonstrate that proteins focused in pH specific bins can be rapidly and efficiently transferred to another location in the same chip without unwanted dilution or dispersive effects. mu IF provides a rapid and versatile option for integrated sample prep or multidimensional analysis, and addresses the glaring proteomic need to isolate trace analytes from high-abundance species in minute volumes of complex samples.

• 出版日期2011-4-15