In order to obtain the identity of sample components, capillary electrophoresis can be directly coupled with mass spectrometers or Surface Enhanced Raman Spectroscopy (SERS). The sensitivity of the technique is attributed to the high intensity of the incident light and the ability to accurately focus the light on the capillary. Laser-induced fluorescence has been used in CE systems with detection limits as low as 10 -18 to 10 -21 mol. The method requires that the light beam be focused on the capillary, which can be difficult for many light sources. The set-up for fluorescence detection in a capillary electrophoresis system can be complicated. This mode of detection offers high sensitivity and improved selectivity for these samples, but cannot be utilized for samples that do not fluoresce. įluorescence detection can also be used in capillary electrophoresis for samples that naturally fluoresce or are chemically modified to contain fluorescent tags. Both of these methods, however, will decrease the resolution of the separation. The capillary tube itself can be expanded at the detection point, creating a "bubble cell" with a longer path length or additional tubing can be added at the detection point as shown in figure 2. To improve the sensitivity, the path length can be increased, though this results in a loss of resolution. According to the Beer-Lambert law, the sensitivity of the detector is proportional to the path length of the cell. The path length of the detection cell in capillary electrophoresis (~ 50 micrometers) is far less than that of a traditional UV cell (~ 1 cm). Bare capillaries can break relatively easily and, as a result, capillaries with transparent coatings are available to increase the stability of the cell window. The portion of the capillary used for UV detection, however, must be optically transparent. In general, capillaries used in capillary electrophoresis are coated with a polymer for increased stability. The use of on-tube detection enables detection of separated analytes with no loss of resolution. In these systems, a section of the capillary itself is used as the detection cell.
The majority of commercial systems use UV or UV-Vis absorbance as their primary mode of detection. Separation by capillary electrophoresis can be detected by several detection devices. Separated chemical compounds appear as peaks with different retention times in an electropherogram. The data is then displayed as an electropherogram, which reports detector response as a function of time. The output of the detector is sent to a data output and handling device such as an integrator or computer. The analytes separate as they migrate due to their electrophoretic mobility, as will be explained, and are detected near the outlet end of the capillary. It is important to note that all ions, positive or negative, are pulled through the capillary in the same direction by electroosmotic flow, as will be explained. The migration of the analytes is then initiated by an electric field that is applied between the source and destination vials and is supplied to the electrodes by the high-voltage power supply. To introduce the sample, the capillary inlet is placed into a vial containing the sample and then returned to the source vial (sample is introduced into the capillary via capillary action, pressure, or siphoning). The source vial, destination vial and capillary are filled with an electrolyte such as an aqueous buffer solution. The system's main components are a sample vial, source and destination vials, a capillary, electrodes, a high-voltage power supply, a detector, and a data output and handling device. A basic schematic of a capillary electrophoresis system is shown in figure 1. The instrumentation needed to perform capillary electrophoresis is relatively simple.
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