A SHEATH FLOW CELL-BASED LASER INDUCED FLUORESCENCE DETECTION SYSTEM WITH PRE-IMAGING SPATIAL FILTERING FOR REDUCED DIMENSION SEPARATION TECHNIQUES

Abstract

The construction, performance, and practical implementation of a rugged device for laser induced fluorescence (LIF) detection in capillary electrophoresis (CE) and liquid chromatography (LC) techniques is described. This device is based on a sheath flow cell (SFC) arrangement with attributes that include ruggedness, simplicity, ease of use, low cost, intra-day reproducibility, and small size. With this design, a pre-imaging mode of spatial filtering is employed to effectively reduce the contributions of both wall scatter and wall fluorescence to the background signal levels. A comparison of the pre-imaging spatial filtering and traditional methods of far-field spatial filtering is performed. This SFC system has been designed specifically for use in the UV spectral region, which can be very advantageous since it broadens the scope of the possible applications of LIF; however, fluorescence of the flow cell walls is particularly problematic in the UV region. The effect of flow cell geometry, slit widths, and detector position on fluorescent background levels are shown. The device has been applied to the fluorescence detection of three dansyl amino acids for both CE and capillary LC techniques. Efficiencies on the order of >105 plates/meter and 104 plates/meter are shown for CE and capillary LC techniques respectively. A comparison is made with CE between on-column and post-column detection demonstrating the latter preserves the integrity of a chromatographic separation.