JPS6258460B2 - - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to an electrophoretic analyzer, and in particular,
The present invention provides an electrophoresis analyzer that can suitably analyze trace components in samples such as urine, blood, and seawater.

For example, when the sample is seawater, seawater contains a large amount of chlorine ion components, and these chloride ion components have high mobility. Chlorine ion components are detected endlessly. In other words, this means that analysis time is wasted.

In view of these circumstances, the inventors of this invention conducted intensive research and found that they divided electrophoresis into two stages and roughly separated the target trace components and chloride ion components in the first stage of electrophoresis. After removing the chlorine ion component that comes out first, the second step of electrophoresis is to precisely separate the target trace components, and the first step of electrophoresis uses a large current in a pretube with a large pipe diameter. We discovered that if the second stage of electrophoresis is performed using a small current in a capillary tube with a small diameter, it is possible to shorten analysis time and stably detect trace components at once. Completed the invention.

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

Reference numeral 1 shown in FIG. 1 is an embodiment of an electrophoretic analyzer according to the present invention.

One end of the constant current power supply circuit 2 is connected to the terminal liquid electrode tank 3, and the other end is connected to the first leading liquid electrode tank 4 and the second leading liquid electrode tank 5 via the switching circuit 16.

These electrode tanks 3, 4, and 5 are connected by a pipe,
A sample injection section 6 and an electric gradient detector 7 are provided in this order in a conduit between the terminal liquid electrode tank 3 and the first leading liquid electrode tank 4.

The pipe line between the sample injection part 6 and the detector 7 includes a pretube 8 with a large pipe diameter (for example, 1 mm) and a capillary tube 9 with a small pipe diameter (for example, 0.3 mm) connected in series via a connecting step part 10. The conduit to the second leading liquid electrode tank 5 is a lead-in tube 12 that branches from the connection stage 10 to the second leading liquid electrode tank 5 . This retraction tube 12 has approximately the same diameter as the pretube 8.

The AD converter 13 converts the supply voltage V ₀ from the constant current power supply circuit 2 into a digital quantity and outputs it to the microcomputer 14 . The microcomputer 14 samples the output of the AD converter 13 at predetermined short time intervals based on a built-in clock. As detailed later, these A-D
The converter 13, the microcomputer 14, and the connecting step 10 constitute passage detection means that outputs an output when the boundary surface of the ion component compartments having different mobilities passes through the connecting step 10.

Further, the microcomputer 14 is an arithmetic control means for controlling the constant current power supply circuit 2, the switching circuit 16, the detector 7, and the passage detection means constituted by itself, and operates as described in detail below.

First, in this electrophoresis analyzer 1, an interface between the terminal liquid and the leading liquid is created in the sample injection part 6 by the usual procedure, and a "blank" is selected from the operation console 15 without injecting a sample, that is, in a blank condition.
Input the command "Start". This command causes the microcomputer 14 to start the constant current power supply circuit 2.
and the switching circuit 16 to apply a constant current between the terminal liquid electrode tank 3 and the first leading liquid electrode tank 4.
I ₁ (e.g. 200-300 μA) is supplied. If the constant current _I1 is maintained as it is, the leading liquid ion component section L and the terminal liquid ion component section T undergo uniform electrophoresis between the electrode vessels 3 and 4, as shown in FIG. At this time, as shown in FIG. 2, the supply voltage V ₀ from the constant current power supply circuit 2 increases, but before and after the rear end boundary surface α of the leading liquid ion component section L passes through the connecting step 10. The rising slopes are markedly different, as shown in a and b. This is because the terminal liquid ion component, which has a conductivity smaller than that of the leading liquid ion component, enters the capillary tube 9.
Therefore, the supply voltage via the A-D converter 13
Microcomputer 1 monitoring V ₀
4 immediately detects the time t ₀ when the interface α passes through the connecting step 10 from the change in the rising slope of the supply voltage V ₀ . Then, the microcomputer 14 memorizes the time t ₀ and controls the constant current power supply circuit 2 to change the supplied current value to a constant current.
I ₂ (e.g. 20-100 μA). One of the reasons for decreasing it is that the electrical conductivity of the terminal liquid ions is low, so if a constant current I ₁ with a large value is maintained, Joule heat becomes excessive, and the supply voltage V ₀ also becomes too high. So the supply voltage
The actual change in V ₀ is as shown in c in Figure 2. Since the leading liquid ion component section L and the terminal liquid ion component section T continue isokinetic electrophoresis under the constant current _I2 , the detector 7 will eventually obtain a signal as shown in FIG. Therefore, the microcomputer 14 detects the time t ₁ when the boundary surface α reaches the detector 7 and stores it. Assuming that the start time of electrophoresis is 0, the above t ₀ is the time τ during which the rear end interface α of the leading liquid ion component section L electrophores from the sample injection port 6 to the connecting stage 10 at a constant current I ₁ The above t _{1 is} equal to the boundary surface α
is the sum of the time τ ₁ for electrophoresis from the connecting stage 10 to the detector 7 at a constant current I ₂ and the above-mentioned τ ₀ .

Next, in this electrophoresis analyzer 1, an interface between the terminal liquid and the leading liquid is created in the sample injection section 6 by a normal procedure, and a sample of 1/n of the predetermined amount is injected therein. Specifically, for example, if the sample is urine and it is normally necessary to inject 1μ in order to detect trace components contained in it, then one-tenth of that amount, 0.1μ, is injected. or inject 1μ of urine diluted 10 times with terminal fluid. Then, from the operation console 15, a command of "1/n start" is input. In response to this command, the microcomputer 14 supplies a constant current I ₁ between the electrode tanks 3 and 4 in the same manner as at the time of "blank start", but after supplying the previously memorized t ₀ time, the current value is changed to the constant current I ₂ Change to The detector 7 obtains a signal as shown in FIG. 3 through this electrophoresis. Here, S indicates an ionic component section of the sample, most of which is a section S' of non-target components with relatively high mobility. There is a very small section S'' of the desired trace component, but this is so small that it is hardly detected. If it is smaller, the result will be as shown in Fig. 3, and if it is the same, the result will be as shown in Fig. 3.The microcomputer 14 calculates the front end boundary surface β of the terminal liquid ion component section T from the output signal of the detector 7. Detect and store the time t ₂ when the current reaches the detector 7. This t ₂ is the time τ ₂ from when the constant current I ₂ starts being supplied until the detector 7 detects the interface β and the τ The microcomputer ₁₄ then calculates the following equation using an appropriate safety factor C (for example, 0.95) of 1 or less that has been input from the console 15 in advance.

(B) τ ₀ = t ₀ (B) τ ₃ = (t ₂ - t ₁ ) x n x I ₂ /I ₁ (C) τ = C x (τ ₀ + τ ₃ ) Next, this electrophoresis analyzer 1, create an interface between the terminal liquid and the leading liquid in the sample injection part 6 according to the usual procedure, inject a predetermined amount of sample (for example, 1 μm of urine) there, and issue a command to "measure and start" from the operation console 15. Enter. Then, the microcomputer 14 controls the constant current power supply circuit 2 and the switching circuit 6 to supply a constant current I ₁ between the terminal liquid electrode tank 3 and the second leading liquid electrode tank 5 for a period of τ. This τ time is a little shorter than the sum of τ ₀ shown in the above calculation formula (a) and τ ₃ shown in (b), but τ ₀ is the time when the rear end boundary surface α of the leading liquid ion component section L is connected. This is the time it takes to reach the stepped portion 10, and _τ3 is the time required for a predetermined amount of the ion component compartment S of the sample to reach and pass through one point when electrophoresed at a constant current _I1 . , means the time until a portion slightly before the rear end of the ionic component section S of the sample reaches the connecting step portion 10. Therefore, the microcomputer 14
After τ time, constant current power supply circuit 2 and switching circuit 1 are connected again.
6 is controlled to supply a constant current _I2 between the terminal liquid electrode tank 3 and the first leading liquid electrode tank 4.
Only the portion mainly containing S'' is precisely separated and detected by the capillary tube 9.

As described above, according to this electrophoresis analyzer 1, the non-target components that make up most of the sample are quickly migrated to the intake tube 12 outside the analysis system using a large current, and the target trace components are made the main component. Since only the portion can be detected by slowly migrating with a small current in the capillary tube 9, analysis time is shortened and analysis can be performed with high precision.

In the above example, a "blank start" was performed to determine τ ₀ and t ₁ , but the conductivity of the ionic component section S' of the non-target component of the sample was lower than that of the leading liquid ionic component section L. If you know in advance that it will be extremely small, you can omit the “blank start” and suddenly start “1/n”.
In this case, since t ₀ and t ₁ have not been set, the microcomputer 14 sets the time at which the passage detection means first outputs an output to t ₀ and detects the leading liquid ions detected by the detector 7. The rear end detection time of component section L is assumed to be _t1.Other operations are the same as in the previous embodiment.

Generally, the electrophoretic analysis apparatus of the present invention is extremely useful for electrophoretic analysis of a sample containing a trace amount of a target component and a large amount of non-target components with higher mobility.

[Brief explanation of the drawing]

Fig. 1 is an explanatory diagram of the configuration of an embodiment of the electrophoretic analyzer of the present invention, Fig. 2 I is a schematic diagram of the ion component compartment at the time of blanking, the same is a diagram showing the temporal change in the supply voltage, and the same is the detection Output signal diagram of the device, Figure 3 I
is a schematic diagram of the ion component compartment during sample injection;
is a diagram of the output signal of the detector. 1... Electrophoresis analyzer, 2... Constant current power supply circuit,
3...Terminal liquid electrode tank, 4...First leading liquid electrode tank, 5...Second leading liquid electrode tank,
6... Sample injection part, 7... Detector, 8... Pretube, 9... Capillary tube, 10... Connecting stage part,
11...Two-stage tube, 12...Retractable tube, 1
3...A-D converter, 14...Microcomputer, 15...Operation console, L...Leading liquid ion component compartment, T...Terminal liquid ion component compartment, α
...the rear end boundary surface of the leading liquid ionic component compartment,
β...front end boundary surface of terminal liquid ionic component compartment,
S...Ionic component compartment of the sample, S'...Ionic component compartment of non-target components in the sample, S''...Ionic component compartment of the target trace component in the sample, 16...Switching circuit.

Claims (1)

[Scope of Claims] 1. A terminal liquid electrode tank is connected to one end of a constant current power supply circuit, and a first leading liquid electrode tank and a second leading liquid electrode tank are connected to the other end via a switching circuit, A sample injection part and a detector are connected in order between the terminal liquid electrode tank and the first leading liquid electrode tank by a pipe line, and the pipe line between the sample injection part and the detector has a large pipe diameter. It is a two-stage tube in which a breech tube and a thin capillary tube are connected in series through a connecting portion, and a lead-in tube with a large pipe diameter is branched from the connecting portion and connected to the second leading liquid electrode tank. and a passage detection means that outputs an output when a boundary surface between ion component compartments having different mobilities passes through the connecting portion, and arithmetic control means, the arithmetic control means being connected to the constant current power supply circuit. By controlling the switching circuit, the detector, and the passage detection means, (i) the time τ ₀ during which the rear end interface of the leading liquid ion component compartment electrophores with a constant current I ₁ from the sample injection port to the connecting portion; Measure and memorize the electrophoresis time τ ₁ from the connecting part to the detector at a constant current I ₂ , (ii) Inject 1/n of the sample amount of the predetermined amount and keep the constant current I ₁
When electrophoresis is performed for τ ₀ hours and then electrophoresis is performed at a constant current I ₂ , the time τ ₂ from when the constant current I ₂ starts to be supplied until the detector detects the front end boundary surface of the terminal liquid ion component compartment is measured. and calculate τ=C×{τ ₀ +(τ ₂ −τ ₁ )×n×I ₂ /I ₁ with C as an appropriate safety factor smaller than 1, (iii) a predetermined amount of When a sample is injected, a constant current I ₁ is supplied between the terminal liquid electrode bath and the second leading liquid electrode bath for a period of τ, and then a constant current I ₂ is supplied between the terminal liquid electrode bath and the first leading liquid electrode bath.
1. An electrophoretic analyzer, characterized in that it supplies an electrophoretic analyzer and performs an electrophoretic analysis.