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Biosensor for detecting cathepsin B as well as detection method and application thereof

A cathepsin and biosensor technology, applied in the field of detection and analysis, can solve problems such as poor sensitivity, and achieve the effects of large dynamic range, high amplification efficiency and high separation efficiency

Active Publication Date: 2020-05-19
SHANDONG NORMAL UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, these methods require the design of complex FRET dye pairs and suffer from poor sensitivity

Method used

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  • Biosensor for detecting cathepsin B as well as detection method and application thereof
  • Biosensor for detecting cathepsin B as well as detection method and application thereof
  • Biosensor for detecting cathepsin B as well as detection method and application thereof

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Experimental program
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Embodiment 1

[0091] medicines and materials. All oligonucleotides and diethylpyrocarbonate (DEPC) treated water were purchased from Takara Biotechnology Co., Ltd. (Dalian, China), and peptide C term -Biotin-KGFRLC-N term Synthesized by Hangzhou Zhongpei Biochemical Co., Ltd. (Hangzhou, Zhejiang, China). Cathepsin B, cathepsin L, cathepsin S, antiprotease and leupeptin were purchased from Sigma-Aldrich (St. Louis, MO, USA), and cathepsin D was purchased from R&D Systems (Minneapolis, MN States, United States) to purchase. Streptavidin magnetic beads, Bst DNA polymerase (large fragment), 10 × ThermoPol reaction buffer (200 mmol tris(hydroxymethyl)aminomethane-hydrochloric acid (Tris-HCl), 100 mmol ammonium sulfate (( NH 4 ) 2 SO 4 ), 100 mmol potassium chloride (KCl), 20 mmol magnesium sulfate (MgSO 4 ), 1% polyethylene glycol octylphenyl ether, pH 8.8), restriction endonuclease (Nt.BstNBI), 10×NEB buffer 3.1 (1000 mmol sodium chloride, 500 mmol tris(hydroxy Methyl)aminomethane-hydro...

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Abstract

The invention provides a biosensor for detecting cathepsin B as well as a detection method and application thereof, and belongs to the technical field of detection analysis. The biosensor at least comprises a polypeptide-DNA conjugate, a DNA template 1 and a DNA template 2. First primer DNA containing an amino acid residue is converted into new primer DNA containing no amino acid residue, the steric hinerance effect in a nucleic acid amplification process can be completely eliminated, and the amplification reaction efficiency can be improved. Meanwhile, proceeding of multiple chain replacementreactions and the circulating digestion function of ribonuclease H can further amplify a fluorescence signal, so that ultra-sensitive detection of the activity of cathepsin B can be realized, and thebiosensor has good practical application values.

Description

technical field [0001] The invention belongs to the technical field of detection and analysis, and in particular relates to a biosensor for detecting cathepsin B, a detection method and an application thereof. Background technique [0002] The information disclosed in this background section is only intended to increase the understanding of the general background of the present invention, and is not necessarily regarded as an acknowledgment or any form of suggestion that the information constitutes the prior art already known to those skilled in the art. [0003] Cysteine ​​cathepsins are a class of proteases located in intracellular lysosomal vesicles that function to mediate bulk dissolution of proteins. Cysteine ​​cathepsins are mainly divided into 11 types of cathepsins B, C, F, H, L, K, O, S, V, W, and X according to different catalytic centers, which play an important role in antigen presentation and bone resorption. effect. Cathepsin B can cleave specific peptide se...

Claims

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Application Information

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IPC IPC(8): C12Q1/37C12Q1/6825
CPCC12Q1/37C12Q1/6825G01N2333/96466C12Q2531/119C12Q2523/308C12Q2563/131C12Q2521/327C12Q2563/107
Inventor 张春阳王子月张程鹏
Owner SHANDONG NORMAL UNIV
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