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Simple two-step isoelectric focusing separation analytic device

A technology of isoelectric focusing and separation analysis, which is applied in the direction of material analysis, measurement device, and material analysis by electromagnetic means, which can solve the problem of hindering the development and application of the two-step isoelectric focusing separation method, cannot provide linear pH gradient, connection To solve problems such as troublesome capillary, to achieve the effect of stable test results, simple structure and convenient operation

Inactive Publication Date: 2006-10-18
TSINGHUA UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, these two migration methods have their own disadvantages: chemical migration, whether using salt or zwitterions, cannot provide a linear pH gradient, which is not conducive to the determination of the isoelectric point (pI) value
However, auxiliary equipment is required for differential pressure migration, the experimental device is more complicated, and it is more troublesome to connect to the capillary.
This hinders the development and application of the two-step isoelectric focusing separation method to a certain extent. Therefore, a simple device is designed to make the migration zone easy to control. certain practical significance

Method used

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  • Simple two-step isoelectric focusing separation analytic device
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Examples

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Embodiment 1

[0021] Example 1: The present invention is used for isoelectric focusing to separate lysozyme and bovine serum albumin.

[0022] The isoelectric focusing separation channel 33 is a dimethylpolysiloxane-coated channel with a film thickness of 0.06 μm, a channel width of 100 μm, a channel length of 5 cm, and an effective length of 4.2 cm. The cathode buffer solution is 20 mM 0.2% MC solution, and the anode buffer solution is 120 mM 0.2% MC solution. The focusing voltage is 1KV; the liquid transported by the micro electroosmotic pump 1 is methanol / water=30 / 70; the driving voltage of the micro electroosmotic pump 1 is 100V. The detection wavelength of the ultraviolet-visible capillary electrophoresis detector on the column is 280nm. A good separation effect was obtained.

Embodiment 2

[0023]Example 2: The present invention is used for isoelectric focusing to separate hemoglobin and cytochrome c. The isoelectric focusing separation channel 33 is a polydimethylsiloxane (PDMS) channel with a channel width of 200 μm, a channel length of 5 cm, and an effective length of 4 cm. Cathode buffer solution is 0.7% NaOH solution, anode buffer solution is 0.1mM H 3 PO 4 solution, the derivatized dye is fluorescein isothiocyanate (FITC). The focusing voltage is 2KV; the liquid transported by the micro electroosmotic pump 1 is methanol / water=40 / 60; the driving voltage of the micro electroosmotic pump 1 is 150V. The detection wavelength of the laser-induced fluorescence detector is 488nm. Good separation.

[0024] In summary, the device of the present invention is simple, and is suitable for the separation and analysis of amphoteric substances such as proteins by the two-step isoelectric focusing method.

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Abstract

The invention relates to a simple two-step isoelectric focusing separation analysis device, which is characterized in that it includes a chip, external high and low voltage DC power supplies and a detection system, and the chip is sequentially provided with micro-electroosmotic pumps, A sampling system, an isoelectric focusing separation system and a waste liquid pool, the high-voltage direct current power supply is loaded on the isoelectric focusing separation system, and the low-voltage direct current power supply is loaded on the micro electroosmotic pump and the sampling system. The present invention adopts the chip as the base body of the device, and through the connection with the external high and low voltage power supply, the micro electroosmotic pump, the sampling system, the isoelectric focusing separation system, the detection system, etc. are all fabricated on the chip, so that the overall device structure is simple , Small size, easy to use. The invention can adjust the output flow rate of the electroosmotic pump by controlling the driving voltage of the electroosmotic pump, and then adjust the speed of the migration focus zone, and is suitable for the separation and analysis of various proteins and other amphoteric substances by the isoelectric focusing two-step method. High efficiency and good detection effect.

Description

technical field [0001] The invention relates to a chip device, in particular to a simple two-step isoelectric focusing separation analysis device suitable for amphoteric substances such as proteins. Background technique [0002] The isoelectric focusing separation method is a mode of capillary electrophoresis separation analysis, which is widely used in the separation of protein mixture samples with different isoelectric points. The isoelectric focusing separation method is divided into two forms: one-step method and two-step method. The one-step method is also called the dynamic focusing method, which uses the joint action of pH gradient and electroosmosis to make the zone migrate in focus or focus in migration. The two-step method is to carry out the focusing process and the migration process in the isoelectric focusing separation separately. The two-step method has higher detection sensitivity, separation efficiency and better reproducibility than the one-step method, s...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N27/453G01N27/447G01N27/26
Inventor 罗国安陈令新刘科辉王义明姚波
Owner TSINGHUA UNIV
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