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Method for preparing protein chip by in-situ synthesis

A technology of protein chip and in-situ synthesis, which is applied in the process of producing decorative surface effects, manufacturing microstructure devices, decorative arts, etc., and can solve the problems of difficult control of experimental conditions, insufficient repeatability of experimental results, and cumbersome manufacturing processes, etc. problems, to avoid the uncertainty of protein conformation and orientation, and to avoid surface non-specific adsorption

Inactive Publication Date: 2012-12-26
WUXI ZHONGMEI YIXIN BIOTECH
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The first is the cost issue. The current protein chip production process is quite cumbersome and complicated, and the signal detection also requires special and expensive equipment (such as seldi-tof-ms); Small changes may cause differences in the final results, and the experimental conditions are not easy to control, making the repeatability of the experimental results relatively insufficient

Method used

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  • Method for preparing protein chip by in-situ synthesis
  • Method for preparing protein chip by in-situ synthesis
  • Method for preparing protein chip by in-situ synthesis

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Example 1: Preparation of aniline derivative monomers with biologically active units

[0030] Dissolve biotin glycol phosphoramidite in anhydrous acetonitrile, then add anhydrous acetonitrile solution of aniline-o-methanol, wherein: the molar ratio of biotin glycol phosphoramidite to aniline-o-methanol is 10:1~1:10 ; Add an appropriate amount of 5-ethylthiotetrazolium (ETT) activator, so that the concentration of the formed activator anhydrous acetonitrile solution is 0.25mol / L; after stirring at room temperature for 30 minutes, carry out vacuum distillation to powder; Add excess oxidation solution, the oxidation solution is formed by dissolving elemental iodine in tetrahydrofuran: water: triethylamine = 8:1:1 mixed solvent, the mass percent concentration of the solution is 4%; after stirring at room temperature for 10 minutes, Reduce to neutrality with sodium thiosulfate, then distill THF under reduced pressure, extract with chloroform, dry the organic layer over anhyd...

Embodiment 2

[0033] Example 2: Preparation of protein chips

[0034] Build microelectrode arrays on individually addressable logic circuit chips (see figure 1 shown), each electrode on this circuit is connected by a switch of a complementary metal oxide semiconductor (CMOS) transistor, by sending an electronic address signal to the common node circuit and then to the SRAM (static random access memory) associated with each electrode body) to turn on the switch; the microelectrode array is placed in a specially designed fluid reactor, and the computer digitally instructs the microelectrode array to respond to digital commands to assemble the conductive polymer; the reaction liquid in the reactor is prepared with The aniline monomer of the biotin substituent is mixed with 1 to 10 times the molar amount of aniline and an appropriate amount of potassium chloride as the electrolyte; under the control of the logic circuit, the pulse is 10 milliseconds on and 10 milliseconds off, at 0.5 to 10 mill...

Embodiment 3

[0037] Embodiment 3: Demonstrate the relation experiment of electric current density and reaction time and the degree of polymerization of conductive polyaniline

[0038] The positive charge is passed on the electrode at different pulse intervals, and the total charge is determined by the integral of the pulse and time. For number 1, the pulse is on for 2 milliseconds and off for 10 milliseconds; for number 2, the pulse is on for 4 milliseconds and off for 10 milliseconds; that is, for each next number, the pulse on time is doubled, and the total time remains unchanged for 60 seconds , the voltage is always maintained at 1.0 volts, as shown in Table 1.

[0039] Table 1 Correspondence between numbers and pulses

[0040] number

[0041] The electrodes were rinsed with deionized water and then blown dry with nitrogen, and then optical measurements were performed. Such as Figure 4 As shown, the number 1 is barely visible, the number 2 is gradually clear, and the num...

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Abstract

In order to solve the problem, which troubles academic circles for a long time, about how to permanently bond protein molecules to the metal actively and fixe the protein molecules on the metallic surface of any detector or any independent addressing electrode in place at one step, the invention provides a method for preparing a protein chip. The method comprises the following steps of: by adopting an in-situ electrochemical combination synthesis method and independent addressing microelectrode array technique, making an aniline derivative monomer containing bioactive units and anilines in solution copolymerized on an electrode in a mode of active free radical polymerization, generating an electrode coverings which is bioactive and electrically conductive on the electrode in situ at one step, and under bionic conditions, making avidins connected with various proteins tightly bonded with the e electrode coverings to prepare the protein chip. The method has the advantages of laying a foundation for an unmarked analysis method, truly making mass production of protein chips possible, providing the possibility for the automatic production of the protein chips at the operation level for the first time, along with high reproducibility.

Description

technical field [0001] The present invention relates to a preparation method of a protein chip, in particular, relates to an in-situ electrochemical combinatorial synthesis (combinatorial synthesis) method and independent addressing microelectrode array technology to make aniline derivatives with biologically active units The monomer is copolymerized with aniline in the solution on the electrode by living free radical polymerization, and a biologically active and conductive electrode covering is directly formed on the electrode in situ in one step, and avidin, which is connected with a variety of proteins, is made under biomimetic conditions. A method for making a protein chip by firmly combining with it. Background technique [0002] Biochip technology is a high-tech developed rapidly in the field of life science since the 1990s. It mainly refers to the construction of a miniature biochemical analysis system on the surface of a solid chip through micromachining and microel...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N33/68G01N27/327B81C1/00
Inventor 王巍
Owner WUXI ZHONGMEI YIXIN BIOTECH
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