Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Palindrome molecular beacon-based biosensor for detecting ATP and fabrication method and application of biosensor

A biosensor and molecular beacon technology, applied in the field of biosensors based on palindromic molecular beacons to detect ATP, can solve the problems of high cost, long detection cycle, low specificity and sensitivity, and achieve low detection limit and fast detection speed , cheap effect

Active Publication Date: 2019-12-24
UNIV OF JINAN
View PDF17 Cites 1 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] In order to solve the problems of relatively low specificity and sensitivity, high cost, and long detection period of the method for detecting ATP in the above prior art, the present invention provides a fluorescent biosensor based on palindromic fragment-mediated bidirectional enzyme repair amplification. for detection of ATP

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Palindrome molecular beacon-based biosensor for detecting ATP and fabrication method and application of biosensor
  • Palindrome molecular beacon-based biosensor for detecting ATP and fabrication method and application of biosensor
  • Palindrome molecular beacon-based biosensor for detecting ATP and fabrication method and application of biosensor

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0044] The preparation method of described fluorescent biosensor comprises the following steps:

[0045] The main steps of the reaction process in homogeneous solution are as follows:

[0046] a. Add AP1 (3 μL, 1 μM), AP2 (3 μL, 1 μM), MB (final concentrations are 1.0 μM, 1.5 μM, 2.0 μM, 2.5 μM, 3.0 μM), Bst DNA polymerase (0.8 U), UDG ( 0.8U), endonuclease IV (0.8U), dNTPs (3 μL), buffer (3 μL) and 3 μL ATP (100 nM) were added to a centrifuge tube, shaken for 30 seconds, and placed in a water bath at 50°C for 90 minutes.

[0047] b. Dilute the solution (30 μL) after the reaction in step a to 100 μL, and detect the fluorescence peak intensity at 518 nm with a fluorometer.

[0048]The excitation wavelength of the fluorescence instrument is set to 486nm, the emission wavelength is 518nm, and the detection range is 500 nm-650 nm to read the change of the fluorescence signal and detect the target object.

[0049] The preparation method of the solution used in the above process: ...

Embodiment 2

[0054] The preparation method of described fluorescent biosensor comprises the following steps:

[0055] The main steps of the reaction process in homogeneous solution are as follows:

[0056] a. AP1 (3 μL, 1 μM), AP2 (3 μL, 1 μM), MB (3 μL, 10 μM), Bst DNA polymerase (0.8 U), UDG (0.2 U, 0.4 U, 0.6 U, 0.8 U, 1.0 U, 1.2U), endonuclease IV (0.8U), dNTPs (3 μL), buffer (3 μL) and 3 μLATP (100 nM) were added to a centrifuge tube, shaken for 30 seconds, and placed in a water bath at 50°C for 90 minutes.

[0057] b. Dilute the solution (30 μL) after the reaction in step a to 100 μL, and detect the fluorescence peak intensity at 518 nm with a fluorometer.

[0058] The excitation wavelength of the fluorescence instrument is set to 486nm, the emission wavelength is 518nm, and the detection range is 500 nm-650 nm to read the change of the fluorescence signal and detect the target object.

[0059] The preparation method of the solution used in the above process:

[0060] 1. Ultrapure...

Embodiment 3

[0064] The preparation method of described fluorescent biosensor comprises the following steps:

[0065] The main steps of the reaction process in homogeneous solution are as follows:

[0066] a. Add AP1 (3 μL, 1 μM), AP2 (3 μL, 1 μM), MB (3 μL, 10 μM), Bst DNA polymerase (0.8 U), UDG (0.8 U), endonuclease IV (0.8 U), dNTPs (3 μL), buffer (3 μL) and 3 μL ATP (100 nM) were added to a centrifuge tube, shaken for 30 s, and placed in a water bath at 35 °C, 40 °C, 45 °C, 50 °C, 55 °C, 60 °C for 90 min.

[0067] b. Dilute the solution (30 μL) after the reaction in step a to 100 μL, and detect the fluorescence peak intensity at 518 nm with a fluorometer.

[0068] The excitation wavelength of the fluorescence instrument is set to 486nm, the emission wavelength is 518nm, and the detection range is 500 nm-650 nm to read the change of the fluorescence signal and detect the target object.

[0069] The preparation method of the solution used in the above process:

[0070] 1. Ultrapure w...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention relates to the technical field of biosensors, in particular to a palindrome molecular beacon-based biosensor for detecting adenosine triphosphate (ATP). The palindrome molecular beacon-based biosensor comprises a palindrome molecular beacon MB, an ATP adaptor (split adaptor probes AP1 and AP2), target object ATP, Bst DNA polymeras, UDG, endonuclease IV and a buffer liquid. Accordingto specificity identification of an aptamer and the target object, fragment tail sequences of the two split adaptors get close to each other, a hairpin structure of the palindrome molecular beacon MBis opened to generate fluorescence, at the moment, a palindrome sequence locked in advance is released, intermolecular hybridization is performed, so that the spontaneous polymerization, repair, internal cut and circulation process is triggered, the amplification of a fluorescent signal is achieved, and an aptamer biosensor is built. The sensor reaction only needs a step, thus, the biosensor has the advantages of rapid detection speed, low cost, low detection limit, high specificity and the like and is simple and convenient to operate.

Description

technical field [0001] The invention relates to the technical field of biosensors, in particular to a biosensor for detecting ATP based on a palindromic molecular beacon, and also relates to a preparation method and application thereof. Background technique [0002] Adenosine triphosphate (ATP), present in all organisms from microorganisms to higher animals and plants, is the main energy source of cells and plays an important role in intracellular metabolic, synthetic and biochemical processes. The content of ATP in the body is not high, and the molar concentration in cells is usually 1-10 mM. Dead cells will not be able to detect ATP, and the ATP metabolism of cancer cells is particularly active. Usually, abnormal ATP concentration is closely related to many diseases, such as hypoglycemia, cardiovascular disease, Parkinson's, tissue hypoxia, etc. Therefore, the rapid and accurate determination of ATP is of great significance for the study of the physiological activity and ...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
IPC IPC(8): G01N21/64
CPCG01N21/6428G01N21/6486G01N2021/6432
Inventor 王玉李莎莎刘素黄加栋徐艺城张儒峰赵一菡瞿晓南孙文玉王业茹
Owner UNIV OF JINAN
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com