JP2940211B2 - Capillary electrophoresis device - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a capillary electrophoresis apparatus for separating and analyzing charged substances such as amino acids, proteins and nucleic acids.

[0002]

2. Description of the Related Art In a capillary electrophoresis apparatus, a substance to be electrophoresed in a capillary is detected by a detector by a voltage applied to both ends of the capillary, and the elution time of each peak is detected by a data processor from the detection signal. The component of each peak is identified from the elution time. Since the mobility of each peak component is determined by the type of the component and the situation in which the component is placed, the retention time is an index for identification as in the case of chromatography. Since the elution time fluctuates, a method has been adopted in which, as in the case of chromatography, correction is performed by taking the ratio of the elution time using a standard substance to cancel the fluctuation of the elution time.

[0003]

Various factors can be considered as causes of the change in the elution time of capillary electrophoresis. Some of the fluctuation factors are affected by both electroosmosis and electrophoresis, and others are only affected by electroosmosis. Since the elution time is represented by the sum of both electroosmosis and electrophoresis, the fluctuation cannot be removed simply by taking the ratio with the elution time of the standard substance. For example, when the temperature of the solution changes, both electroosmosis and electrophoresis fluctuate at the same rate for each substance, so that the fluctuation can be removed. However, when the inner surface state of the capillary changes due to dirt or the like, the factor of the electroosmotic flow changes independently, and the change cannot be removed by the conventional method using the standard substance. SUMMARY OF THE INVENTION An object of the present invention is to provide a capillary electrophoresis apparatus capable of performing correct identification by removing fluctuations in elution time due to electroosmosis.

[0004]

In the present invention, the elution time of a neutral substance is measured to determine the electroosmotic flow rate. Using this, the portion that contributes to electroosmosis is removed from the elution time of each ion component, and the elution time when the electroosmosis flow rate is assumed to be 0 is determined.

The present invention will be described with reference to FIG. The elution time detecting means 2 detects an elution time of each peak from a detection signal of a substance which electrophoreses in the capillary by a voltage applied to both ends of the capillary. The electroosmotic flow rate calculating means 4 calculates the electroosmotic flow rate from the elution time of the peak of the neutral substance.
The electrophoretic velocity calculating means 6 determines the electrophoretic velocity from the elution time of each peak detected by the elution time detecting means 2 and the electroosmotic flow velocity calculated by the electroosmotic flow velocity calculating means 4. The elution time correction means 8 determines the elution time of each peak corrected by setting the electroosmotic flow rate to 0.

[0006]

In the capillary electrophoresis apparatus, the velocity Va of the ionic species a in the capillary is represented by Va = Vo + Vea. Here, Vo is the speed of the electroosmotic flow (electroosmosis), Vea is the electrophoresis speed of the ionic species a, and Vo and Vea are respectively expressed as Vo = μo · E Vea = μea · E. μo is the mobility of electroosmosis, μea is the mobility of ionic species a, and E is the strength of the electric field.

Assuming that the distance from the sample injection section to the detector is L, the time (elution time) ta until the ion species a in the sample is detected is expressed as follows. ta = L / Va = L / {(μo + μea) · E} Assuming that the elution time of the neutral substance or the added neutral substance in the sample is ts, Vo = L / ts. Since (Vo + Vea) · ta = L for the elution time ta of the ion species a, Vea = L / ta−Vo is obtained from this. Assuming that the electroosmotic flow rate is 0, the relative elution time ta 'can be obtained by ta' = L / Vea. The relative elution time is determined for each ion species, and normal correction using a standard substance is performed using this as the elution time.

FIG. 2 is a flowchart showing the procedure from the detection of the elution time of each peak to the calculation of the relative elution time. When the elution time of each peak is detected, the elution time of the neutral component is selected therefrom to determine the electroosmotic flow velocity Vo, and the electrophoretic velocity V of each peak component is determined using this Vo.
a, Vb,... Next, relative elution times ta ′, tb ′,... Are determined for each peak component using the electrophoretic velocities Va, Vb,. Time. afterwards,
Identification is performed using the standard substance and the relative elution time.

[0009]

FIG. 3 shows an embodiment. Both ends of the capillary 12 are immersed in buffer solutions in the reservoirs 14 and 16, respectively, and a migration voltage is applied to both buffer solutions from a high voltage power supply 18. A sample is injected into the end of the capillary 12 on the reservoir 14 side, and the sample is electrophoresed in the direction of the reservoir 16. On the reservoir 16 side, a detector 20 for detecting components migrating in the capillary 12 is provided. From a signal detected by the detector 20, the elution time of each component is detected by a data processing device 22, Relative elution time of each peak component based on the elution time of the substance
Required by 2.

The elution time detecting means 2, the electroosmotic flow velocity calculating means 4, the electrophoretic velocity calculating means 6 and the elution time correcting means 8 in FIG.
The data processing device 22 is realized by, for example, a personal computer or a microcomputer.

[0011]

According to the present invention, the flow rate of the electroosmotic flow is obtained from the elution time of the neutral substance, and the relative elution time excluding the influence of the electroosmotic flow is obtained from the detected elution time of each peak component. For example, it is possible to remove a variation factor of the elution time due to, for example, a change in the inner surface of the capillary, and to more accurately correct the variation of the elution time due to a change in temperature, thereby performing accurate identification.

[Brief description of the drawings]

FIG. 1 is a block diagram illustrating the present invention.

FIG. 2 is a flowchart showing the operation of the present invention.

FIG. 3 is a schematic diagram showing one embodiment.

[Explanation of symbols]

 2 Elution time detecting means 4 Electroosmotic flow velocity calculating means 6 Electrophoretic velocity calculating means 8 Elution time correcting means 12 Capillary 20 Detector 22 Data processing device

Claims (1)

(57) [Claims] 1. A capillary electrophoresis apparatus for detecting a substance to be electrophoresed in a capillary by a voltage applied to both ends of the capillary with a detector, and measuring the elution time of each peak by a data processor from the detection signal. In addition to the elution time detecting means for detecting the elution time, the data processing device is an electroosmotic flow velocity calculating means for obtaining the electroosmotic flow velocity from the elution time of the neutral substance peak, and the elution time detecting means for each peak. An electrophoresis speed calculating means for obtaining an electrophoretic speed from the detected elution time and the electroosmotic flow speed obtained by the electroosmotic flow speed calculating means, and an elution time for obtaining an elution time of each peak corrected by setting the electroosmotic flow speed to 0. A capillary electrophoresis device comprising a correction unit.