JP4085514B2 - Electrophoresis chip - Google Patents

Description

[0001]
BACKGROUND OF THE INVENTION
The present invention relates to an electrophoresis member used for electrophoresis for analyzing trace amounts of proteins, nucleic acids, drugs, etc. at high speed and with high separation, and more specifically, electrophoresis is performed in a separation channel formed inside a transparent plate member. The present invention relates to an electrophoresis chip used in a microchip electrophoresis apparatus.
[0002]
[Prior art]
In the case of analyzing a very small amount of protein or nucleic acid, an electrophoresis apparatus has been conventionally used, and a typical example is a capillary electrophoresis apparatus. A capillary electrophoresis apparatus is a device in which an electrophoresis buffer is filled in a glass capillary having an inner diameter of 100 μm or less, a sample is introduced into one end, and then a high voltage is applied between both ends to develop an analyte in the capillary. is there. Since the capillary has a large surface area relative to the volume, that is, the cooling efficiency is high, it is possible to apply a high voltage, and it is possible to analyze a trace amount sample such as DNA with high speed and high resolution.
[0003]
Since the capillary has a thin outer diameter of about 100 to 500 μm and easily breaks, it has a problem that it is not easy to handle at the time of replacement of the capillary to be performed by the user. Therefore, DJ Harrison et al./ Anal. Chim. Acta 283 (1993) 361-366 shows a form that can be expected to speed up analysis and reduce the size of the device instead of capillaries that are complicated to handle. In addition, an electrophoresis chip (referred to as a microchip) formed by joining two substrates has been proposed. An example of the microchip is shown in FIG.
[0004]
The microchip 1 includes a pair of transparent plate-like inorganic materials (for example, glass, quartz, silicon, etc.) or substrates 1a and 1b made of plastic, and is used for migration that crosses the surface of one substrate 1b by a micromachining technique. Capillary grooves 3 and 5 are formed, and through holes 7 are provided as reservoirs at positions corresponding to the ends of the grooves 3 and 5 on the other substrate 1a. The microchip 1 is used in a state where both the substrates 1a and 1b are overlapped and bonded as shown in FIG.
[0005]
When electrophoresis is performed using the microchip 1, the electrophoresis buffer is filled from any one of the reservoirs 7 into the grooves 3 and 5 by, for example, pressure feeding using a syringe prior to analysis. Next, the electrophoresis buffer filled in the reservoir 7 at one end of the shorter groove (sample introduction flow path) 3 is replaced with the sample and attached to the electrophoresis apparatus. In addition, the loading of the electrophoresis buffer and the filling of the sample may be performed after the microchip 1 is mounted on the apparatus, and depending on the apparatus, the filling of the electrophoresis buffer and the filling of the sample may be performed automatically.
[0006]
Next, a predetermined voltage is applied to each reservoir 7, the sample is migrated into the groove 3, and is guided to the intersection 9 between the grooves 3 and 5. Next, the voltage applied to each reservoir 7 is switched, and the sample existing at the intersection 9 is injected into the groove 5 by the voltage between the reservoirs 7 and 7 at both ends of the longer groove (separation channel) 5. Then, the sample is separated in the groove 5. By arranging a detector at an appropriate position in the groove 5, the sample separated by electrophoresis is detected. Detection is carried out by means such as absorptiometry, fluorometry, electrochemical or electrical conductivity.
The analysis conditions such as the flow path design of the microchip 1 and the composition of the electrophoresis buffer differ depending on the application and sample.
[0007]
[Problems to be solved by the invention]
When performing microchip electrophoresis, it is necessary to fill the microchip with the electrophoresis buffer for each analysis and to keep the liquid level of the electrophoresis buffer filled in each reservoir constant. Performing this operation manually using a syringe or the like is complicated for the operator. Moreover, even if this operation is automated, there is a problem that the apparatus becomes large and takes a long time.
[0008]
When optically detecting a separated sample by light absorption or fluorescence, the surface of the microchip on the side irradiated with light should be cleaned so that no light is absorbed or scattered by reagents, scratches, dust, etc. Need to keep. This leads to the need to consider the analysis environment and operator education, and cannot be said to have good operability.
Therefore, an object of the present invention is to provide a microchip that can start analysis immediately.
[0009]
[Means for Solving the Problems]
The microchip of the present invention is an electrophoresis chip in which a channel is formed inside a transparent plate member, and a hole reaching the channel is formed at a position corresponding to the channel on one surface of the transparent plate member. The flow path and the hole are filled with a predetermined migration buffer, and at least the hole on one surface of the transparent plate member is covered with a film member.
[0010]
A microchip on which a flow path design suitable for an analysis application is formed is prepared, and a flow buffer and a reservoir (hole) of the microchip are preliminarily filled with an electrophoresis buffer that is an optimal analysis condition. Furthermore, at least the reservoir is covered with a film member. The microchip is stored so that the loaded electrophoresis buffer is not denatured, and the analysis can be started immediately by removing the film member and attaching it to the apparatus immediately before use. In addition, when the sample can be injected through the film member with a nozzle, it is not necessary to peel off the film member.
[0011]
DETAILED DESCRIPTION OF THE INVENTION
Furthermore, if the position corresponding to the flow path on the surface of the microchip is also protected by the film member, the detection position can be prevented from being contaminated with a reagent or dust by peeling the film member immediately before use. Optical detection can be performed with good reproducibility.
When an electrode pattern for applying an electrophoretic voltage to the hole is formed on one surface of the transparent plate member, the electrode pattern is also preferably covered with a film member. As a result, the electrode pattern can be protected until just before use, and malfunction due to damage to the electrode pattern can be suppressed.
[0012]
【Example】
2A and 2B are diagrams illustrating an embodiment, in which FIG. 2A is a perspective view, and FIG. 2B is a cross-sectional view taken along line AA ′ in FIG.
A separation channel 5 is formed inside the microchip 1, and a sample introduction channel 3 orthogonal to the separation channel 5 is formed on one end side of the separation channel 5. A reservoir 7 is provided at a position corresponding to the end of the separation channel 5 and the sample introduction channel 3 on one surface of the microchip 1. When the microchip 1 is used for analysis, the separated sample is detected at a detection position 11 surrounded by a one-dot chain line on the other end side of the separation channel 5.
[0013]
The sample introduction channel 3, the separation channel 5, and the reservoir 7 are filled with an electrophoresis buffer. The electrophoresis buffer is sent from one of the reservoirs 7 using a syringe, for example, and filled so that the flow paths 3 and 5 and the other reservoirs 7 are completely filled. The running buffer is thoroughly degassed before filling.
The migration buffer is not particularly limited, and any migration buffer such as an inorganic ionic buffer such as phosphoric acid or boric acid, or an organic polymer such as methylcellulose or hydroxymethylcellulose may be used.
[0014]
A plastic film 13 is bonded to the surface of the microchip 1 on the side where the reservoir 7 is formed by adhesion or heat fusion. The film 13 seals the reservoir 7, covers the flow paths 3, 5, and is stuck so that no adhesive or debris of the film 13 remains at the detection position 11. When an adhesive is used, the adhesive is not applied to the reservoir 7 portion. A non-adhesive portion 13 a is provided at one corner of the film 13 so that the film 13 can be easily peeled off.
[0015]
The material of the film 13 is not particularly limited, and polyethylene, polypropylene, polypropylene copolymer, polystyrene, polyvinyl chloride, thermoplastic elastomer, polycarbonate, polyethylene terephthalate copolymer, tetrafluoroethylene, fluorinated ethylene propylene, poly Any material can be used as long as it can seal the electrophoresis buffer and does not cause chemical reaction with the electrophoresis buffer, such as vinylidene fluoride.
[0016]
When the microchip 1 is used, the microchip 1 is mounted on the apparatus immediately before the analysis, and the film 13 is peeled off from the non-adhesive portion 13a (see FIG. 2A). Thereafter, the electrophoresis buffer contained in any one of the reservoirs 7 in the sample introduction channel 3 is removed, and instead, the sample is injected into the reservoir 7 by the same volume. Then, a predetermined electrophoresis voltage is applied to each reservoir 7 to start analysis.
Furthermore, in electrophoresis, the film 13 may be passed through the reservoir 7 to pierce the electrode, and the electrophoresis may be performed without removing the film 13.
[0017]
FIG. 3 is a perspective view showing another embodiment.
Four electrode patterns 15 for applying an electrophoresis voltage to each reservoir 7 are formed on the surface of the microchip 1 on the side where the reservoir 7 is formed. Each electrode pattern 15 is formed to reach the inner wall of one of the reservoirs 7. The electrode pattern 14 is made of gold, for example, and is formed by vapor deposition.
[0018]
A plastic film 17 is attached to the surface of the microchip 1 on the side where the electrode pattern 15 is formed by adhesion or heat fusion. The film 17 seals the reservoir 7, covers the sample introduction channel and the separation channel (not shown), and is attached so as to also cover the electrode pattern 15. A corner portion of the film 17 protrudes from the microchip 1 and is formed so that the film 17 is easily peeled off from the corner portion.
In this embodiment, in addition to the effects of the embodiment of FIG. 2, the electrode pattern 15 can be prevented from being damaged.
[0019]
【The invention's effect】
In the microchip of the present invention, a predetermined migration buffer is filled in the flow path and the reservoir, and at least the reservoir on one surface of the microchip is covered with a film member, so that the migration buffer is filled immediately before the analysis. This saves time and effort, so analysis time can be shortened and analysis can be started immediately. Furthermore, since the functions to be provided in the microchip electrophoresis apparatus can be simplified, the reliability of the apparatus can be improved, and miniaturization and portability can be achieved.
In addition, if the position corresponding to the flow path on one surface of the microchip is also covered with the film member, contamination of the microchip surface at the detection position can be prevented.
In addition, when an electrode pattern for applying an electrophoretic voltage to the reservoir is formed on one surface of the microchip, the electrode pattern can be used with the microchip if the electrode pattern is also covered with a film member. It is possible to protect until just before, and to suppress malfunction due to damage to the electrode pattern.
[Brief description of the drawings]
FIGS. 1A and 1B are diagrams illustrating an example of a microchip, in which FIG. 1A is a top view of one substrate, FIG. 1B is a top view of the other substrate, and FIG. FIG.
2A and 2B are diagrams illustrating an embodiment, in which FIG. 2A is a perspective view, and FIG. 2B is a cross-sectional view taken along line AA ′ in FIG.
FIG. 3 is a perspective view illustrating another embodiment.
[Explanation of symbols]
DESCRIPTION OF SYMBOLS 1 Microchip 3 Sample introduction flow path 5 Separation flow path 7 Reservoir 11 Detection position 13 Plastic film 13a Non-adhesion part

Claims (2)

In the electrophoresis chip in which a flow path is formed inside the transparent plate-shaped member and a hole reaching the flow path is formed at a position corresponding to the flow path on one surface of the transparent plate-shaped member,
The flow path and the hole are filled with a predetermined electrophoresis buffer,
An electrophoresis chip, wherein a position corresponding to at least the hole and the flow path on the one surface of the transparent plate member is covered with a film member. On the one surface of the transparent plate-shaped member, the electrode pattern is formed for applying a migration voltage into the hole, the electrophoresis chip according to claim 1, which is covered by the electrode patterns film member .

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