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System for adsorbing, separating and detecting ultra-drop target protein

A target protein, ultra-micro-volume technology, applied in material separation, measurement devices, selective adsorption, etc., can solve problems such as insufficient elution of chip microbeads, small amount of modified samples, and affecting the specificity of chip microbeads

Inactive Publication Date: 2008-08-20
重庆黄嘉同怡科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, this type of microbead chip has its inherent disadvantages: 1. Chip specificity, which is related to the surface modification of the chip microbeads and the modifiers. The surface modification process of the chip microbeads is not good or the distance between the modifiers is not suitable. Non-specific adsorption on the surface of microbeads, and the specificity of modifiers directly determine the specificity of the chip
2. Chip capacity, the surface area of ​​chip microbeads is limited, so the number of modifiers modified is certain, therefore, chip capacity has always been an important issue for microbead chips
3. The combination of the chip microbeads and the sample is not sufficient. The combination of the chip microbeads and the molecules to be tested is a planar combination, and the amount of sample required for the chip microbeads is very small, so it cannot be guaranteed that the chip microbeads are fully combined with the sample. Like the liquid, the two items can be fully mixed; 4. The chip microbeads are not fully eluted. The chip is a solid plane, and the elution cannot completely wash the microbeads, which may cause incomplete washing of interfering substances and affect the specificity of the chip microbeads; 5. , The amount of modified sample is small, and the ability to detect low-abundance target substances is limited

Method used

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  • System for adsorbing, separating and detecting ultra-drop target protein
  • System for adsorbing, separating and detecting ultra-drop target protein
  • System for adsorbing, separating and detecting ultra-drop target protein

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0081] Embodiment one: the preparation of microbead

[0082] 1. Chitosan Microbeads

[0083] Chitosan (chitosan), chemical name (1,4)-A-amino-A-deoxy-B-D-glucan, is a copolymer of N-deacetylglucosamine, because chitosan and its derivatives are free of Toxicity, biodegradability and good biocompatibility, it is widely used as immobilized carrier of enzymes and cells.

[0084] The following is the preparation process of chitosan microbeads: weigh chitosan 9, dissolve it in 300mL of 2% acetic acid solution, stir continuously for 2 hours to fully dissolve, remove insoluble matter by suction filtration, add Tween-80 10mL, oil Phase dispersion medium, porogen ethyl acetate, emulsifier and polymer surface modifier, fully stirred and reacted at 50°C for 30 minutes, raised the temperature to 60°C, added formaldehyde solution, reacted for 30 minutes, then added 2.0ml of glutaraldehyde, Adjust the pH to 9.0, raise the temperature to 80°C, and react for 60 minutes. Suction filter, wash ...

Embodiment 2

[0099] Embodiment 2: Surface functional group modification of microbeads

[0100] 1. Surface affinity modification process of chitosan microbeads

[0101] Add appropriate amount of deionized water to chitosan microbeads, activate with epichlorohydrin, add EDC and heparin, and stir at 4°C for 24 hours to obtain surface affinity-modified microbeads.

[0102] The long-chain alkyl group of chitosan microbeads is used as the hydrophobic part, and the sulfate group is used as the hydrophilic part to synthesize N-octyl-O-sulfate chitosan (OCS1) to obtain microbeads with surface hydrophilic modification.

[0103] 2. Surface ion exchange modification process of cellulose microbeads

[0104] Add the washed cellulose microbeads to a solution containing 0.1mol / L DEAE hydrochloride and 0.5mol / L NaOH, and keep stirring at 60°C for 10 hours to make it cross-link-DEAE weakly basic anion exchange group group.

[0105] 3. Metal chelation affinity modification of microbeads

[0106] Add micr...

Embodiment 3

[0107] Embodiment three: the loading of the separation filler in the hollow tube

[0108] Put the fine steel wire mesh or wire mesh with smaller diameter into the upper end hole, and it is located in the middle or lower part of the hollow tube, and then put the separation packing. According to the number of samples to be separated, adjust the density of the separation packing. Finally, a fine steel wire mesh or wire mesh with a larger diameter is installed to fix the separation packing.

[0109] Or, in the process of preparing microbeads, close or plug the upper end hole of the hollow tube until the upper limit position of the separation filler, then insert the hollow tube upside down into the microbead solution, adjust the position of the separation filler in the hollow tube, and the separation filler After being fixed in the hollow tube, the closure or blockage is removed from the upper end of the hollow tube to obtain a hollow tube containing separated packing without a cy...

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Abstract

The invention provides a system for absorbing, separating, purifying and testing trace target protein, which includes the separating filling, the hollow tube for fixing separating filling and combining with solution and the absorbing solution. The invention applies the micro bead of protein micro bead chip in the field of chromatography, elution and separation, that is used as the carrier of chromatography separation column, creatively changes the regular chromatography separation column environment into the environment similar to removing liquid instrument tip, in order to obtain application effects of adding sample, absorbing, cleaning, eluting, condensing, identifying in the same environment, and at the same time the high specific application effect of protein micro bead chip .

Description

technical field [0001] The invention relates to the separation of ultra-trace target proteins in biological liquid samples in proteomics and molecular biology, and specifically provides a system for separating and detecting ultra-trace target proteins in biological liquids and a method for using the same. technical background [0002] With the rapid development of life science research, the separation method of target protein in biological samples has attracted more and more attention of scientific researchers. Scientists have found that with the emergence of many sensitive and accurate methods for protein detection, such as fluorescent quantitative PCR instruments and mass spectrometers, the problem that currently limits protein research is how to effectively process early samples. [0003] There are also many methods for processing target proteins in biological samples. The most common ones are chromatographic elution separation methods (chromatographic methods), such as ...

Claims

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Application Information

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IPC IPC(8): G01N33/68G01N30/00B01D15/08B01J20/281
Inventor 于中连马庆伟赵洪斌吕芳吕萍萍
Owner 重庆黄嘉同怡科技有限公司
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