Method for realizing protein separation detection by using PVP/CdS quanta dot modified electrode

A technology for protein separation and electrode modification, which is applied in the field of electrodes, can solve the problems of limitation and slow electron transfer speed, and achieve the effects of low detection limit, high sensitivity and good reproducibility

Inactive Publication Date: 2006-02-08
EAST CHINA NORMAL UNIV
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  • Abstract
  • Description
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Problems solved by technology

The HPLC-ECD system composed of thin-layer ECD and HPLC has been successfully applied to neurotransmitters such as catecholamines and their metabolites such as dopamine and norepinephrine, and electroactive amino acids such as cysteine ​​and glutathione. detection of biologically active small molecules, but so far there has been no report that the thin-layer ECD has been applied to the HPLC-ECD system to achieve protein separation and detection
The main reason is that the electroactive center of the protein is embedded in the polypeptide chain due to the huge three-dimensional structure of the protein, and the adsorption and denaturation of the protein on the surface of the glassy carbon electrode will cause the passivation of the electrode surface, so the electrons of the protein on the glassy carbon electrode The transfer speed is very slow and does not produce an effective current response, which limits the application of glassy carbon electrodes in direct electrochemical studies of proteins

Method used

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  • Method for realizing protein separation detection by using PVP/CdS quanta dot modified electrode
  • Method for realizing protein separation detection by using PVP/CdS quanta dot modified electrode
  • Method for realizing protein separation detection by using PVP/CdS quanta dot modified electrode

Examples

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Embodiment 1

[0034] Example 1 One of the methods for protein separation and detection using PVP / CdS quantum dot modified electrodes

[0035] The protein chromatographic column is VARIAN Microsorb 300-5C4 chromatographic column of American VARIAN Company.

[0036] In the second step, the flow rate of the mobile phase is 1.0mL / min, the mobile phase changes at a rate of 2% / min within the gradient range of 20% to 60% A, and the working voltage applied to the ECD by the electrochemical workstation is -0.3V , pre-run for 10 minutes. In the third step, configure the standard concentration as 1.0×10 -8 mol / L~1.0×10 -4 mol / L solution of three proteins: hemoglobin, myoglobin and cytochrome c, the peak times of the chromatographic peaks of the three proteins: hemoglobin, myoglobin and cytochrome c are 12.49min, 9.02min and 6.33min respectively , and its concentration-current working curves are respectively y=1.0841x+2×10 -8 , y=1.1537x+1×10 -8 and y=1.2451x+1×10 -8 , where y and x are the respo...

Embodiment 2

[0037] Example 2 The second method of using PVP / CdS quantum dot modified electrode to realize protein separation and detection

[0038] In the second step, except that the following conditions are different from Example 1, other conditions are the same as Example 1: the flow rate of the mobile phase is 2.0mL / min, and the mobile phase is in the range of 20%~60%A gradient at 4% / Min speed change, pre-run 12min. In the third step, the peak times of the chromatographic peaks of hemoglobin, myoglobin and cytochrome c are 6.25min, 4.51min and 3.17min respectively; Three chromatographic peaks appeared at 3 and 6.25 minutes respectively, and the peak currents were 0.73μA, 0.51μA and 0.89μA, respectively. By comparing with the standard peak time measured in the third step, it can be determined that the proteins to be tested corresponding to the three chromatographic peaks are cytochrome c, myoglobin and hemoglobin respectively. By comparing with the working curve measured in the thir...

Embodiment 3

[0039] Example 3 Method 3 of Using PVP / CdS Quantum Dot Modified Electrode to Realize Protein Separation and Detection

[0040] In the second step, except that the following conditions are different from Example 1, other conditions are the same as in Example 1: the flow rate of the mobile phase is 5.0mL / min, and the mobile phase is in the range of 20%~60%A gradient at 10% / min. Min speed change, pre-run 15min. In the third step, the peak times of hemoglobin, myoglobin and cytochrome c were 2.49min, 1.80min and 1.23min, respectively. In the fourth step, after the mixed sample 3 was injected, three chromatographic peaks appeared at 1.23min, 1.80min and 2.49min respectively, and the peak current values ​​were 98nA, 75nA and 84nA, respectively. By comparing with the standard peak time measured in the third step, it is determined that the proteins to be tested corresponding to the three chromatographic peaks are cytochrome c, myoglobin and hemoglobin respectively. By comparing with...

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Abstract

The invention relates to a method for using PVP / CdS quantum point modification electrode to achieve protein dividing detecting in the field of nanometer modification electrode and biology detecting. The method must be carried out in the HPLC-ECD architecture, which is formed by the protein chromatographic column with protein, the HPLC and working electrode as PVP / CdS quantum point modification electrode. It uses the HPLC technique to divide the protein; the HOLC pump drives the traveling phase and the detected liquid crossed the protein chromatographic column to enter into the ECD directly and uses PVP / CdS quantum point modification electrode to do electrochemist to the divided protein.

Description

technical field [0001] The invention relates to a method for protein separation and detection using a PVP / CdS quantum dot modified electrode, to be more precise, relates to a method for using a PVP / CdS quantum dot modified electrode as a protein high performance liquid chromatography (HPLC)-electrochemical detector ( The electrode of the ECD in the ECD) system, the method for realizing protein separation and detection, belongs to the technical field of nano-modified electrodes and biological detection. Background technique [0002] Proteins are information macromolecules that play key roles in life processes. After the completion of the human genome sequence ahead of schedule, proteomics research has attracted great attention from life science workers. Finding fast and efficient means of protein separation and detection has become a hot spot of concern. [0003] HPLC is one of the most important methods in protein separation. The protein detection metho...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N30/64G01N30/02
Inventor 金利通刘梅川张莉程欲晓张文鲜跃仲施国跃
Owner EAST CHINA NORMAL UNIV
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