A laser-induced fluorescence imaging system for isoelectric focusing

作者：Das C; Xia Z; Stoyanov A; Fan ZH^*

来源：Instrumentation Science & Technology, 2005, 33(4): 379-389.

DOI：10.1081/CI-200063695

• laser-induced fluorescence
• imaging system
• isoelectric focusing
• proteins

This paper describes a laser-induced fluorescence (LIF) imaging system for detecting proteins separated by isoelectric focusing (IEF). IEF was carried out in either microfabricated glass devices or in capillaries. Compared to the single-point-detection used in capillary IEF (CIEF), the imaging approach eliminates the need for mobilizing the focused proteins to the detector, thereby reducing proteins' de-focusing during mobilization and improving separation resolution. The LIF imaging system consists of a laser; its diameter is first increased through a beam expander and subsequently the laser beam is focused into a line using a cylindrical lens. The resultant laser line is used to image an entire channel or capillary in which protein separation take place. The fluorescence emission is collected with a cooled, scientific grade charge-coupled device (CCD) camera. The detection limit was determined using a series of concentrations of fluorescein solutions. The imaging system was demonstrated by isoelectric focusing of proteins, including Green Fluorescent Protein, B-phycoerythrin, and R-phycoerythrin. In addition, we can obtain temporal profiles of protein separation, providing useful information about the dynamic behavior of protein migration during IEF.

• 出版日期2005-8