Hydrostatic pressure, sample introduction

Sample introduction To introduce the sample, the left end of the capillary is dipped into a sample vial with several centimeters hydrostatic pressure for a fixed number of seconds. This will force a small volume of liquid sample into the end of the capillary. Another method of sample introduction is to dip the end of the capillary into the sample vial and turn on the power for a few seconds. Sample ions thus flow into the system by electrical migration. 

Separations should initially be attempted with the capillary thermostatted at close to ambient temperature. The capillary temperature can be increased on most commercial CE units to as high as 60 C without substantially increasing current with most buffers. When this is done using the same applied voltage, decreased buffer viscosity leads to an increase in analyte electrophoretic mobility, thus, decreasing separation times. Also, it is important to note that, when sample introduction is hydrostatic (same pressure/vacuum and time), increased capillary temperature will lead to an increase in the injected sample volume as a result of decreased buffer viscosity. " Sensitivity may not necessarily be increased. However, Undesirable effects include concurrent changes in buffer pH, band broadening due to increased diffusion, and possible thermal denaturation of the sample. 

Figure 4.85 Basic components of a CE system (1) fused-silica capillary (2) electrolyte vessels with electrodes (3) syringe-to-capillary adaptor (replaced in commercial instruments by pressure or vacuum-driven rinse) (4) sample vial raised to a level necessary for sample introduction by hydrostatic pressure (5) regulated high-voltage power supply (6) detector (7) data acquisition device.

The introduction of samples into capillaries by means of differential pressure (hydrostatic injection) has become the most popular method in capillary electrophoresis. For sample introduction, the sample vial is raised to a defined level above the detection reservoir for a specified time frequently about 5 s. To terminate the injection, the end of the capillary is removed from the sample and replaced into the vial containing the background electrolyte. 

Fig. 5.1 The relaxation of (a) the square radius of gyration, S2, and (b) the hydrostatic pressure at 500 K following the introduction of excluded volume in a sample of a melt composed of 640 chains each with 100 monomers of model PE II polyethylene (see Section 5.3). Note the expanded vertical scale. The simulations were at constant volume 20 bars pressure discrepancy would be equivalent to a density discrepancy at constant pressure of about 0.1%. S2 values are shown relative to those determined assuming complete screening of long-range interactions (see text). The density is 0.70 g cm". These results were obtained with a Fujitsu APIOOO massively parallel computer.

The introduction of the samples onto the capillary column can be carried out by either displacement techniques or electrokinetic migration. Three methods of displacement or hydrostatic injection are available a) direct injection, or pressure b) gravity flow, or siphoning and c) suction. The electrokinetic injection method arose from findings that electroosmosis act like a pump (80). Both methods have advantages and disadvantages. For example, a bias has been reported in electrokinetically injected... 

The sample, typically a few nanoliters, can be introduced into the capillary by hydrostatic injection (gravity, pressure, or vacuum) or by electromigration. The sample volume should generally be less than 2% of the total capillary length. For gravity introduction, the capillary sample end is dipped into the sample (which may be as small as 5 fiL) and raised for a short predetermined time to allow sample to flow into the capillary. Or, it is inserted into a pressurized vial to force sample into the capillary. Or it is drawn in by suction from the other end of the capillary. After injection, the sample vial is replaced with a buffer reservoir. Alternatively, the sample end is immersed in the sample solution and a relatively low voltage is ap-phed for a few seconds, for example, 2000 V for 10 s. This injects the small volume of sample by electroosmosis. 

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