JPS635700B2 - - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to an electrophoresis apparatus, and an object of the present invention is to provide an electrophoresis apparatus that can accurately divide the ion component zones of an electrophoresed sample and take out all of them.

As an apparatus for extracting the ionic component zone of an electrophoresed sample, the applicant of the present application has proposed, for example, an apparatus disclosed in Japanese Patent Publication No. 31542/1983. This device adds a target fraction separation channel and a target fraction extrusion liquid inflow channel to the migration tube at the rear of the detector of a normal electrophoresis analyzer, and injects the extrusion liquid into the migration tube. Then, the target portion is pushed out into the separation channel and taken out. However, with this device,
The preparative range becomes uncertain because it is extruded with liquid,
It was difficult to accurately divide and extract the ionic component zone.

On the other hand, as an apparatus capable of accurately dividing electrophoresed ion component zones, there is, for example, an apparatus described in Japanese Patent Application Laid-Open No. 53-3892 by the applicant of the present application. This is a conventional electrophoresis analyzer in which a part of the migration tube at the rear of the detector is replaced with a communication hole drilled in a sliding plate, and when the target part enters the communication hole, the sliding plate is closed. The ionic component zones can be precisely divided by sliding movement. However, this device is not intended for extracting the target part;
Therefore, the configuration was not such that the target portion could be taken out.

Furthermore, all of the conventional devices mentioned above use ordinary detectors to detect the location of the target part, so there is a limit to their detection accuracy, and in reality, even with the latter device, it is difficult to perform high-precision segmentation. It was hot.

The inventor of the present invention conducted extensive research in view of the above circumstances, and as a result, completed a device of the present invention that can solve the problems of the conventional device described above.

That is, the present invention provides a terminal liquid electrode tank,
A sample injection section, a switching conduit section having a switching conduit, a leading liquid electrode tank, a migration conduit connecting these in order, a power supply part whose output ends are connected to the terminal liquid electrode tank and the leading liquid electrode tank, respectively; an arrival detection section that integrates the electrophoresis current supplied by the power supply section over time and detects that the ionic component section of the sample has reached the switching channel section based on the integrated value, and is provided in the switching channel section. a leading liquid injection part, a separation part, and a connection switching means for switching and connecting the switching pipe of the switching pipe part to the electrophoresis pipe, the leading liquid injection part, or the separation part based on the arrival detection means; An electrophoresis device comprising:

The main features of the apparatus of this invention are (i) when the target part enters the switching pipe, the switching pipe is separated from the migration pipe and connected to the separation section to take out the target part; (ii) The entry of the target part into the switching pipe is detected based on the integrated value of the electrophoretic current, and (iii) The leading liquid is injected into the switching pipe after the target part is taken out. The reason is that it can be switched and connected to the electrophoresis tube again later.

Hereinafter, the present invention will be explained in detail based on embodiments shown in the drawings. Note that this invention is not limited to this.

1 shown in FIG. 1 is an embodiment of the electrophoresis apparatus of the present invention. Output end 2 of high voltage power supply circuit 2
A terminal liquid electrode tank 3 and a leading liquid electrode tank 4 are connected to a and 2b, respectively, and a sample injection part 5 and a switching pipe part 6 are connected in order between the electrode tanks 3 and 4 through a migration pipe 7. connected.

The switching pipe section 6 consists of two disc-shaped fixed flanges 8, 8' and a disc-shaped sliding body 9 rotatably and pivotally supported between the fixed flanges 8, 8'. Switching pipes 10, 10', 10'' are bored in the sliding body part equidistant from the rotating shaft (not shown).
At three locations on the fixing flanges 8, 8' that can be aligned with 0'', migration tube connecting parts 11, 11', leading liquid injection parts 12, 12', and separation parts 13, 13' are provided. When the moving body 9 is rotated, the switching pipes 10, 10', 10'' are connected to the connecting parts 11, 1.
1', leading liquid injection parts 12, 12', and separation parts 13, 13'.

A leading liquid inlet 12 is connected to a leading liquid tank 15 via a valve 14.
2' is connected to the drainage channel 16.

The separation section 13 is connected to a nitrogen gas cylinder 18 via a valve 17, and 13' is connected to a separation capillary tube 19. Also, 1 of the preparative section
A switching pipe line 1 is provided with a light source 20 on the 3 side, a light receiving section 21 on the 13' side, and connected to the separation sections 13 and 13'.
It is now possible to measure UV absorption of samples in 0".

The electrophoretic current supplied from the high voltage power supply circuit 2 is transmitted to the microcomputer 2 via the current detection circuit 22.
3 is entered. The current detection circuit 22 and the microcomputer 23 constitute an arrival detection means, which detects to what position in the electrophoresis channel 7 the ion component zone of the sample has electrophoresed. That is, as is well known, the amount of electricity supplied during electrophoresis is directly proportional to the migration distance of one ion component zone. Therefore, by knowing the amount of electricity, the migration distance can be determined. The microcomputer 23 integrates the output of the current detection circuit 22, that is, the electrophoretic current, to obtain the above-mentioned quantity of electricity, and thereby detects the electrophoretic position of the ion component section. Now, if we pay attention to the ionic component zone of the leading liquid, the relationship between the migration distance and the amount of electricity depends only on the composition of the leading liquid. However, since the rear end of the ionic component zone of the leading liquid is the front end of the ionic component zone of the sample,
The migration distance of the front end of the ionic component zone of the sample can also be determined from the relationship between the migration distance of the ionic component zone of the leading liquid and the amount of electricity. In other words, as long as the leading liquid is the same, the migration distance of the ionic component zone of the sample can be accurately determined from the electrical quantity using the same judgment conditions, and there is no need to consider the composition or amount of the sample. become. For this purpose, in this apparatus 1, the composition of the leading liquid and the electricity required for the rear end of the ion component zone of the leading liquid to migrate from the sample injection part 5 to the switching pipe outlet 28 when the leading liquid is used are determined in advance. The quantity Qi and the quantity of electricity qi required for migration from the switching pipe inlet 27 to the outlet 28 are stored in the microcomputer 23 in association with each other. How to obtain these Qi and qi will be explained later.

Reference numeral 24 denotes a motor controlled by the microcomputer 23, which rotates the drive gear 25 to rotate and slide the sliding body 9. That is, the microcomputer 23, the motor 24
and the drive gear 25 are connected to the switching pipes 10, 1
0' and 10'' are connected to the electrophoresis pipe 7, the leading liquid injection parts 12 and 12', and the fractionating parts 13 and 13'.

Reference numeral 26 is an operation console that interacts with the microcomputer 23.

Next, the operation of this device 1 will be explained. In an initial state, the switching pipes 10, 10', 10'' are the migration pipe 7, the leading liquid injection part 12, 12',
It is assumed that it is in a position connected to the fractionating sections 13, 13'. First, the composition of the reading liquid is input into the microcomputer 23 via the console 26.
Then, the microcomputer 23 reads the corresponding electric quantities, for example, Q ₁ and q ₁ from the memory and sets them internally. Next, create an interface between the terminal liquid and the leading liquid in the sample injection section 5 using the usual procedure, inject the sample, and then
Input the start command from 6. With this command, the microcomputer 23 controls the high voltage power supply circuit 2.
is controlled to supply electrophoresis current and start electrophoresis. While electrophoresis is being carried out,
The valve 14 is operated to inject leading fluid into the switching line 10'. The electrophoresis current is fed back to a microcomputer 23 by a current detection circuit 22. Microcomputer 23
integrates this electrophoretic current over time, and
The quantity of electricity Q obtained by integration and the quantity of electricity Q ₁
and stops the electrophoresis current when they match. When the quantity of electricity Q matches the quantity of electricity _Q1 , it is when the rear end of the ionic component zone of the leading liquid has reached the switching pipe outlet 26, as described above, that is, when the front end of the ionic component zone of the sample has reached the This is when the switching pipe 10 is entered.

Therefore, the microcomputer 23
4 to rotate and slide the sliding body 9, and switch the switching pipe 1.
0, 10', 10'' are separated into fractionating sections 13, 13', respectively.
Electrophoresis pipe 7, leading liquid injection part 12, 12'
Switch to connect. The front end portion of the ionic component zone of the sample cut out by the switching pipe 10 and transported to the fractionating sections 13, 13' has its UV absorption measured by a UV absorption detector, and then is processed by a microcomputer using a valve. 17 is activated and supplied under pressure to the preparative capillary tube by nitrogen gas,
taken outside. On the other hand, the switching pipe 10'' connected to the leading liquid injection parts 12 and 12' is injected with the leading liquid in the same manner as described above. 2 to supply electrophoresis current.The ionic component zone of the sample remaining in the pipe line 7 on the side of the sample injection part 5 from the switching pipe part 6 is filled with the leading liquid in the switching pipe line 10'. The microcomputer 23 integrates the electrophoresis current at this time over time, compares the integrated quantity of electricity q with the quantity of electricity _q1 , and when they match, starts electrophoresis again. When the quantity of electricity q matches the quantity of electricity q ₁ , it is when the rear end of the ionic component zone of the leading liquid has moved from the switching pipe inlet 27 to the outlet 28, that is, when the remaining sample This is when the front end portion of the ionic component zone enters the switching conduit 10'.

The following operation is the same as that described above, so the explanation will be omitted, but in the end, the ion component zones of the sample are divided one after another and all are taken out. Since the extracted parts are separated by nitrogen gas, they remain independent within the capillary tube 19, and are lined up in the capillary tube 19 in the order in which they were extracted. Each of these parts is used by being broken off along with the capillary tube 19, collected into a sample bottle or the like, or fixed to some kind of support.

Now, how to find Q ₁ and q ₁ , this is done as follows.

That is, first, a large amount of a UV-absorbing substance is injected as a sample using the usual method, and then appropriate Q _{1 ′} and q _{1 ′} , which are sufficiently smaller than the empirically predicted Q ₁ and q ₁ , are set on the operating console 26. is input to the microcomputer 23, and an electric quantity measurement start command is input. Since the microcomputer 23 initially operates in exactly the same manner as described above, it first calculates the UV absorption value of the portion fractionated when the integrated quantity of electricity Q is Q ₁ '.
U ₁ is obtained, and then the UV absorption value U ₂ of the fraction fractionated after q ₁ is obtained. Q ₁ ′ is the actual
Since it is smaller than Q ₁ , U ₁ is usually the UV absorption value of only the leading liquid and is substantially 0. However, since the ionic component zone of the sample gradually advances, when the electrical quantity Q reaches, say, Q ₄ ', it becomes 0.
A UV absorption value U ₄ that is not measured is measured. more than a certain value
After the UV absorption value is measured, the microcomputer 23 continues to operate while multiplying q ₁ ' by a natural number (that is, lengthening the separation interval by q ₁ minute). Therefore, the rate at which the ionic component zone of the sample enters the switching pipe gradually increases.
The UV absorption value gradually increases. Shown in Figure 2
U ₅ , U ₆ , and U ₇ show this situation. However, if the preparative separation interval exceeds q ₁ , the entire switching pipe becomes the ionic component zone of the sample, so the UV
The absorption value is a constant maximum value. U ₈ shown in Figure 2,
U ₉ shows this situation.

As shown in principle in FIG. 3, the microcomputer 23 calculates q ₁ from the increase curve α and the maximum value β of the UV absorption value with respect to the amount of electricity, and also calculates the increase curve α and the UV absorption value U ₄ From this, calculate the amount of electricity R until just before the front end of the sample ion component zone enters the switching pipe, and add that R and _q1 to obtain _Q1 .
In this way, q ₁ and Q ₁ are found.

As can be understood from the above explanation, according to the electrophoresis apparatus of the present invention, it is possible to accurately divide the ion component zone of the electrophoresed sample and take out all the ion component zones, and the capacity of the switching pipe can be reduced to If it is made sufficiently small compared to the zone, the target zone can be extracted almost purely. Furthermore, the position of the extracted portion relative to the entire ion component zone is extremely clear. Note that if a detector is provided in the electrophoresis channel, it can also be used as a normal electrophoresis analyzer.

[Brief explanation of the drawing]

Fig. 1 is an explanatory diagram of the configuration of an embodiment of the electrophoresis apparatus of the present invention, Fig. 2 is a graph of UV absorption to explain the operation of the apparatus shown in Fig. 1, and Fig. 3 is similar to Fig. 1. FIG. 3 is an explanatory diagram of the principle of operation of the device shown in FIG. 1... Electrophoresis device, 2... High voltage power supply circuit,
3...Terminal liquid electrode tank, 4...Leading liquid electrode tank, 5...Sample injection port, 6...Switching pipe section, 7...Migration pipe line, 8, 8'...Fixed flange,
9...Sliding body, 10, 10', 10''...Switching pipe line, 12, 12'...Leading liquid injection part, 1
3, 13'... preparative section, 22... migration current detection section,
23...Microcomputer, 24...Motor, 25...Drive gear, 26...Operation console.

Claims (1)

[Scope of Claims] 1. A terminal liquid electrode tank, a sample injection part, a switching pipe section having a switching pipe, a leading liquid electrode tank, a migration pipe connecting these in order, the terminal liquid electrode tank and the leading liquid electrode tank. A power supply unit with an output end connected to each of the power supply units, and an arrival unit that integrates the electrophoresis current supplied by the power supply unit over time and detects that the ionic component section of the sample has reached the switching pipe section based on the integrated value. Based on the detection section, the leading liquid injection section and the separation section provided in the switching pipe section, and the arrival detection means, the switching pipe of the switching pipe section is connected to the migration pipe, the leading liquid injection section, or the switching pipe section. An electrophoresis device comprising a connection switching means for switching connection to the separation section. 2. The switching pipe section is composed of fixed flanges fixed to the migration pipe on the sample injection part side and the migration pipe on the terminal liquid electrode tank side, respectively, and a sliding body that slides between the fixed flanges, 2. The apparatus according to claim 1, wherein a switching conduit is bored through the sliding body, and a leading liquid injection part and a separating part are provided in the sliding flange. 3. The apparatus according to claim 1 or 2, wherein the separation section is provided with a detector. 4. The apparatus according to any one of claims 1 to 3, wherein the separation section comprises a separation flow path that can be connected to one end of the switching pipe and a push-out gas supply flow path that can be connected to the other end. . 5. Claims 1 to 4, wherein the arrival detection section comprises a migration current detection circuit, an integration circuit for the output of the migration current detection circuit, and a comparison circuit that compares the output of the integration circuit with a predetermined threshold value. Apparatus according to any of paragraphs.