JP3097199B2 - 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 method for separating and analyzing trace components in a liquid.
Related to multi-functionalization.

[0002]

2. Description of the Related Art Capillary electrophoresis has recently attracted particular attention as a separation and analysis technique having the advantages of high-performance liquid chromatography (HPLC) and gel electrophoresis.

[0003] From the inner diameter of about 25um filled buffer to 2
A 50 um capillary is used as the separation medium. A sample is introduced from one end, in which the sample is moved to the other end while being separated by electrophoresis. A detector for detecting passage of the sample component is installed at an appropriate position in the moving direction, and a separation pattern is recorded.

[0004] For example, analytical chemistry
61, 292A-303A (1989) (Analytic
al Chemistry, 61 , 292A-303A (1989)) and Analytical Chemistry 61, 1186-1194 (1).
989) (Analytical Chemistry, 61 , 1186-1194 (198
9)).

[0005] A single capillary is fixedly used in the apparatus.

[0006]

In capillary electrophoresis, as described above, a capillary having a small inner diameter is used as a separation medium, and a sample is introduced from one end and developed into the other. Small components tend to remain in the capillaries and affect the measurement of the next sample.

In order to obtain reproducible measurement results, it is essential to wash the inside of the capillary for each measurement. However, if analysis is to be performed using a single capillary tube, the following steps are required until the washing is completed. Since measurement of the sample cannot be started, there is a limitation in improving the throughput (the number of measurements per unit time). Also,
If the washing is not performed, effects such as a decrease in reproducibility appear and the life of the capillary is significantly impaired.

In order to analyze various symmetries of measurement, it is necessary to change the developing solution (electrolyte solution, buffer) for each sample according to the characteristics of the object to be measured and the separation technique used, and also to sufficiently exchange the internal solution of the capillary. is necessary.

Further, in order to ensure high resolution, it is necessary to use a capillary tube having a modified surface or a capillary tube filled with a gel for each object to be measured. It takes time and manpower.

[0010]

According to the present invention, a plurality of thin tubes are provided.
And separating the sample introduced into the capillary by electrophoresis.
Voltage applying means for applying a voltage so that
Having a detector for detecting the sample,
Said washed when at least one is being washed
Sample introduced into at least one capillary other than the capillary
Are separated. Or multiple tubules
And separating the sample introduced into the capillary by electrophoresis.
Voltage applying means for applying a voltage so that
Having a detector for detecting the sample,
At least two have internal diameter, length, inner surface treatment or
Or, it is configured so that the properties of the filler and the like are different. There
Swims a plurality of capillaries and a sample introduced into the capillaries.
Voltage applying means for applying a voltage so as to be separated by movement
Having a detector for detecting the separated sample,
Capillary tube comprises multiple dedicated capillaries with different measurement targets
The configuration is as follows. Alternatively, a plurality of capillaries and the capillaries
Apply voltage so that the introduced sample is separated by electrophoresis.
Means for applying voltage, and detecting the separated sample.
At least one of the plurality of capillaries.
Are stored in a container, and the thin tube stored in the container is transferred.
It is configured to be moved and exchanged by a moving mechanism. is there
Alternatively, a plurality of thin tubes and a sample introduced into the thin tubes are electrically
Voltage applying means for applying a voltage so as to separate by electrophoresis
Having a detector for detecting the separated sample,
A plurality of thin tubes are housed in one container, and are installed in the container.
The placed capillary is moved and replaced by the moving mechanism.
Be configured as follows. Alternatively, a plurality of thin tubes and the thin tube
Voltage is applied to separate the input sample by electrophoresis.
Means for applying voltage and detecting the separated sample
A circle having a detector, wherein the plurality of thin tubes have a rotation mechanism
It is installed on the panel, and contacts with the detection unit provided on the rotating shaft.
The connection is switched by rotating the rotating disk.
You. Alternatively, insert a capillary between the light source and the light detector.
Light emitted from the light source passes through the capillary
In the position where the maximum value of the transmitted light intensity is given.
By moving the capillary or light source and photodetector
In order to determine the appropriate position of the capillary.

[0011]

[Function] Since a plurality of thin tubes are installed in the device, high speed
Labor saving by measurement or automation. In particular, since the other capillaries can be washed in parallel even during the sample separation analysis, the measurement efficiency does not decrease even if the washing time is sufficient.
If this can be automatically performed by the device, speeding up and labor saving can be achieved.

If a capillary tube with a different surface treatment or a capillary tube filled with gel or the like is installed in the apparatus, it can be automatically replaced in accordance with a wide variety of objects to be measured.

[0013]

FIG. 1 shows the principle of capillary electrophoresis. The capillary 1 is filled with the buffer 2 and both ends are placed in the electrolytic cell 3. When a sample is introduced into one end of a capillary and a voltage 4 is applied to both ends, the capillary moves toward the other end while being separated according to the mobility of each component in the sample. Detector 6 in the middle of the capillary
And analyze by detecting the passage of each component. For detection, absorbance measurement is often used. Usually, a washing solution or buffer is flowed into the capillary tube for each measurement to wash the inside. That is, the analysis operation is as shown in FIG. 2, and a waiting time for washing is generated for each measurement.

FIG. 3 shows the principle of the present invention. In FIG. 3a,
A method will be described in which a plurality of separation capillaries 7, 7 ', 7 "and a plurality of detectors 8, 8', 8" following them are prepared and exchanged and used at the same time. They can be integrated and put into one box for use. FIG. 3b shows the separation capillaries 9, 9 ',
A method of preparing a plurality of 9 ″ and one set of the detection unit 10 and connecting the capillary to be used and the detection unit by the flow path switching device 11 provided between the separation capillary and the detection unit will be described. Since a plurality of capillaries can be used efficiently, no time is wasted due to cleaning as shown in Fig. 4. Next, a specific example will be described.
m, a fused quartz capillary 12 having an outer diameter of 0.4 mm is housed in a box-shaped housing 13, and both ends thereof are provided with an inlet port 14, respectively.
It was fixed to the outlet port 15. Window 16 for detection in part of the box
Was provided. Since the fused quartz capillary was covered with a resin, the coating of the capillary at this portion was peeled off and removed. This detector was placed between a light source provided outside and a photodiode so that absorbance could be measured. This whole is called a capillary assembly 30.

An example of an apparatus using the capillary assembly 30 will be described with reference to FIGS. The description will be made in the order of the analysis operation. Capillary assembly 30,3
0 ′, 30 ″, 30 ′ ″... Are set on the tray 31.
The samples are put into sample cups 32, 32 ',. The capillary assembly 3 to be used from the tray 31 according to the measurement conditions input to the controller.
Select 0 and move to the analysis unit 34. An optical reading device is mounted on the moving mechanism 45, and identifies the capillary assembly. Each capillary assembly 30,3
At 0 ', 30 ",..., A classification code and a unique code are printed as bar codes so that information such as the inner diameter and length of the capillary, surface treatment, packing, and the number of times of use can be referred to. If magnetic recording or semiconductor memory is used, more detailed information can be recorded, which is useful.

The capillary assembly 30 is fixed by a retainer 36 along a fixing guide 35. The position of the detector 40 is adjusted by the method described below, the level of the recorder is adjusted to 0, and then the sample is introduced into the injection port 38 using the autosampler 37. Perform electrophoresis and start analysis. After the analysis, the used capillary assembly 3
“0” is moved to the cleaning unit 39. In the cleaning unit 39, the cleaning liquid is injected under pressure from the inlet port 14 or the outlet port 15. After the cleaning is completed, the tray is returned to the tray 31. At the time of washing, the analysis of the next sample is performed in parallel. Therefore,
The total analysis time is reduced by about half, and the efficiency can be greatly improved.

By providing two or more cleaning positions, the cleaning time can be extended. If the analysis time and washing time are known in advance, the computer can optimize the order of the measurement sample and the capillary used so that the total required time is the shortest, and can automatically change the order and perform the measurement. It becomes possible.

In accordance with the movement of the capillary assembly,
It was necessary to perform automatic position correction of the detector 40. FIG. 8 shows the principle of the position correcting mechanism 46. The side of the capillary 12 is masked with a material that does not transmit light, and the light source 4
Light from 2 is incident. When the capillary 12 is located at the center of the light beam 43, the transmitted light intensity I becomes highest, so that
While detecting the transmitted light intensity I with the photodiode 44,
The appropriate position may be obtained by slightly moving the capillary assembly 30 or the detector 40. Thereafter, the transmitted light intensity is set to 100% (that is, the absorbance is set to 0), and the absorbance is measured.

For sample injection, injection ports 18, 18 'as shown in FIG. 9 were used.

The structure a is a two-way cock. A sample is injected from the injection port 18 and the measuring section 19 is filled with the sample. Then, the cock 20 is rotated and connected to the migration path 21 to reduce the migration voltage. Apply. The drain portion 22 is made thin to reduce leakage of the sample.

In another structure (b), the operation is performed by operating a valve without a rotating part. Open the valve 24 and load the sample into the inlet 1
From 8 ', it is implanted into the drain portion 22'. At this time, the sample enters the part 25 in the migration path 21 '. After closing the valve 24, an electrophoresis voltage is applied.

Another example will be described with reference to FIG.

The capillaries 51, 51 ', 51 ",... Are arranged radially on a turntable 50, and a switching valve 52 is installed at the center. Only the capillary 51 installed in one direction has a center. The capillaries 51 ', 51 ",... Arranged in the other direction are connected to the drain, which is connected to the detection unit 53 placed on the axis. The switching valve 52 has its shaft side fixed to the main body of the apparatus.
The connection direction is switched.

The sample and the buffer are separate turntables 5
4 and rotated as needed.

The operation will be described in the order of measurement.
Sample cups 55 and 5 in which samples are placed on turntable 54
5 ',... And buffers 56, 56', 56 "are prepared.
The turntable 50 and the turntable 54 are rotated to predetermined positions. The turntable 54 is lifted up, and the capillary 51 is inserted into the sample. Lower electrode 57,
Insert into the sample. The sample is introduced into the capillary 51 by applying a voltage 58. The voltage is lowered, the electrode 57 is raised, the turntable 54 is lowered, and then rotated to the position of the buffer 56. The turntable 54 is lifted again, the electrodes are lowered, and a voltage is applied. The sample is electrophoresed while being separated into each component. The change in concentration is monitored by the detector 53. When the analysis is completed, reduce the voltage,
The electrode is raised, and the turntable 54 is lowered. The turntable 50 is rotated, and the used capillary 51 is moved to the cleaning position. There are a plurality of washing positions, and at the same time, the next capillary 51 ″ to be used is placed at the analysis position. The analysis of the next sample is performed in the same manner.

Washing is performed by sending a washing solution 59 to the capillary by a pump 60.

All operations can be controlled automatically by a computer.

Care must be taken when handling, especially storing, the capillaries. Gel-filled capillaries are vulnerable to drying and must be prevented. FIG. 12 shows an example of a capillary assembly for facilitating handling. The entrance port 14 ' , the exit port 15 ' and the detection window 16 are covered and protected by the slide cover 70 as shown in FIG. As shown in FIG. 12, the inlet port 14 'and the outlet port 15' may be arranged on the side of the housing 13 '.
An identification label 66 is provided, and various information is recorded.

[0029]

According to the present invention, high-speed measurement or automation
Is possible and labor can be saved.

[Brief description of the drawings]

FIG. 1 is a principle diagram of capillary electrophoresis.

FIG. 2 is a diagram showing a flow of analysis by a conventional apparatus.

FIG. 3 is a diagram showing an outline of the present invention.

FIG. 4 is a diagram showing a flow of analysis according to the present invention.

FIG. 5 is a diagram illustrating an example of a capillary assembly.

FIG. 6 is a diagram illustrating an example of a device configuration.

FIG. 7 is a diagram showing details of an analysis unit.

FIG. 8 is a diagram illustrating the principle of a position adjusting mechanism of the detector.

FIG. 9 is a diagram illustrating a method for introducing a sample.

FIG. 10 is a diagram showing another device configuration.

FIG. 11 is a diagram showing details of a switching valve.

FIG. 12 is a view showing an example of a capillary assembly with a cover.

[Explanation of symbols]

DESCRIPTION OF SYMBOLS 1 ... Capillary tube, 2 ... Buffer, 3 ... Electrolyzer, 4 ... High voltage power supply, 5 ... Electrode, 6 ... Detector, 7 ... Separation capillary, 8
... Detection part, 9 ... Separation capillary, 10 ... Detector, 11
... flow path switching device, 12 ... capillary, 13 ... housing, 14 ... inlet port, 15 ... outlet port, 16 ...
Detection window, 18 injection port, 19 measuring unit, 20 cock, 21 electrophoresis path, 22 drain, 24 valve, 2
5: part of migration path filled with sample, 30: capillary assembly, 31: tray, 32: sample cup, 33
... sample tray, 34 ... analysis unit, 35 ... fixed guide, 36
… Hold, 37… Autosampler, 38… Injection port,
39 cleaning unit, 40 detector, 41 mask, 42 light source, 43 light flux, 44 photodiode, 45 moving mechanism, 46 position adjusting mechanism, 50 turntable, 5
DESCRIPTION OF SYMBOLS 1 ... Capillary, 52 ... Switching valve, 53 ... Detector, 54 ... Turntable, 55 ... Sample cup, 56 ...
Buffer, 57: Electrode, 58: High voltage power supply, 59: Cleaning liquid, 60: Pump, 61: Waste liquid container, 70: Slide cover, 71: Identification label.

Claims (4)

(57) [Claims] A plurality of thin tubes; a voltage applying means for applying a voltage so as to separate a sample introduced into the thin tubes by electrophoresis; and a detector for detecting the separated sample. When at least one of the capillaries has been washed, at least one other than the washed capillaries
A capillary electrophoresis apparatus characterized in that a sample introduced into one capillary is separated. 2. A sample is introduced into a plurality of thin tubes, and the sample is charged.
Separation by electrophoresis and detection of the separated sample
And while electrophoresis is taking place, at least one
Characterized in that the capillary is washed and subjected to electrophoresis.
Capillary electrophoresis method. 3. A sample is introduced into a plurality of thin tubes, and the sample is charged.
Separation by electrophoresis and detection of the separated sample
And place at least one capillary in the washing position after electrophoresis
Moving and moving other tubules to the electrophoresis position
Capillary electrophoresis method. 4. The gel according to claim 3, wherein the gel is stored during storage of the capillary.
The outlet of the filled capillary tube and the detection window are protected by a cover.
A capillary electrophoresis method, characterized in that: