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Method for testing ultratrace DNA by dual-amplifying technique and electrochemical quartz crystal microbalance

A dual-amplification, quartz crystal technology, applied in electrochemical variables of materials, biochemical equipment and methods, measuring devices, etc. Effect

Inactive Publication Date: 2004-12-15
WUHAN UNIV +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although QCM is a sensitive mass detector that can detect sub-nanogram mass changes, in actual detection, the concentration of DNA is far below the range of direct detection by QCM, especially as a biological tool for detecting ultra-trace changes. sensor, its sensitivity can not meet the requirements

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0011] Example 1: Adriamycin (ADM) modified nanomagnetic microspheres were used to confirm the formation of targeted double-stranded DNA molecules and perform quality amplification to detect ultra-trace target DNA.

[0012] 1. Preparation of doxorubicin-modified magnetic microspheres: preparation 2.0×10 -6 The aqueous solution of mol / L doxorubicin is used to prepare the cross-linking reaction between the amino group and the aldehyde group. After adding 0.02g aldehyde group magnetic microspheres (available commercially available products), the mixture is placed at 4°C and stirred for reaction 4 After hours, the microspheres were washed 3 times with double distilled water on the magnetic stand, and 0.5 mL of double distilled water was added and stored at 4°C for later use.

[0013] 2. Self-assembled α-lipoic acid monolayer film: Before using the quartz crystal, immerse it in anhydrous methanol solution for 30 minutes, then ultrasonically clean it for 1 minute, then wash it with dist...

Embodiment 2

[0018] Example 2: Actinomycin D (ActD) modified nano-magnetic microspheres were used to confirm the formation of targeted double-stranded DNA molecules and to amplify the quality to detect ultra-trace target DNA.

[0019] Preparation of Actinomycin D-modified Nano Magnetic Microspheres: Preparation 1.0×10 -6 The acetone solution of mol / L actinomycin D is used to prepare the cross-linking reaction between the amino group and the aldehyde group. After adding 0.02g aldehyde group magnetic microspheres, the mixture is stirred and reacted at 4°C for 4 hours. The ball was cleaned 3 times with acetone on a magnetic stand, 0.5 mL of acetone was added, and stored at 4°C for later use. The remaining steps are the same as in Example 1.

Embodiment 3

[0020] Example 3: Albumin nanospheres modified with mitoxantrone were used to confirm and amplify the formation of targeted double-stranded DNA molecules to detect ultra-trace target DNA.

[0021] Preparation of mitoxantrone modified albumin nanospheres: preparation 2.0×10 -6 The aqueous solution of mol / L mitoxantrone is used to prepare the cross-linking reaction between the amino group and the aldehyde group. After adding 0.02g aldehyde-based albumin microspheres, the mixture is stirred and reacted at 4°C for 4 hours, 12000 rpm Centrifuge / min, wash 3 times with double-distilled water, add 0.5 mL of double-distilled water, and store at 4°C for later use. The remaining steps are the same as in Example 1.

[0022] The present invention can also be labeled with doxorubicin drugs such as doxorubicin, epirubicin, zorubicin, and other anthracycline antitumor drugs such as bisantrene, or other drugs or biological reagents that can selectively interact with double-stranded DNA. The nano...

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PUM

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Abstract

A method for detecting ultratrace DNA by electrochemical quartz crystal microbalance and double amplifying technique includes such steps as plating a thin Au or Ag film or quartz crystal, selectively adsorbing DNA by quartz crystal oscillator, molecular hybridizing of DNA to obtain dual-chain DNa, amplifying the detected signals in mass and electrochemical areas by the nano microbead marked by chemical or biologic reagent which can selectively act on the dual chains of DNA and has electrochemical activity, and measuring the mass signal and electrochemical signal. Its advantages are high selectivity and high sensitivity.

Description

Technical field [0001] The invention relates to a method for detecting ultra-trace DNA with a double-amplification technology electrochemical quartz crystal microbalance. Background technique [0002] Quartz crystal microbalance (QCM) is also called piezoelectric sensor, and its theoretical basis is based on piezoelectric effect. The principle of the piezoelectric DNA biosensor is: first plating a thin layer of gold film or silver film on the quartz wafer, and then trying to make the quartz crystal oscillator selectively adsorb DNA, through DNA molecular hybridization, to another DNA containing complementary base sequence Recognize, combine into double-stranded DNA, and use frequency changes to reflect hybridization. Since QCM can have a frequency response to increasing the nanogram-level mass on the surface, it is a very sensitive quality detection device, suitable for detecting the quality change of single-stranded DNA (ssDNA) before and after hybridization, so it can be applie...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68G01N27/30G01N27/327G01N33/50
Inventor 张盛龙丁虹胡先明王海涛罗顺德
Owner WUHAN UNIV
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