Thrombin detection method based on splitter adapter and water-soluble conjugated polymer
A conjugated polymer and detection method technology, applied in the field of fluorescent detection of thrombin, can solve the problems of cumbersome operation, high background of blank samples, affecting detection accuracy and sensitivity, etc., and achieve the effect of rapid method and high sensitivity detection
- Summary
- Abstract
- Description
- Claims
- Application Information
AI Technical Summary
Problems solved by technology
Method used
Image
Examples
Embodiment 1
[0025] 1) Mix dye-labeled aptamer fragment 1, aptamer fragment 2 and water-soluble conjugated polymer at a molar ratio of 1:1:5 to form an aptamer / polymer complex, and then add it to the buffer solution In this method, the fluorescence detection is carried out with 404 nm as the excitation wavelength, the fluorescence emission band of the complex is recorded, and the ratio I of the fluorescence intensity at 525 nm to 440 nm is calculated 525 / I 440 ; Aptamer fragment 1 and aptamer fragment 2 are specific single nucleotide sequences, the 5' end of aptamer fragment 1 is labeled with fluorescein isothiocyanate, and the labeling method is: 5'-6-FITC.
[0026] 2) Add aptamer fragment 1 and aptamer fragment 2 and the test sample containing thrombin into the binding solution and place it at 37°C for 40 minutes to form a thrombin / aptamer complex.
[0027] 3) Adding a water-soluble conjugated polymer to the above complex, thrombin induces and combines with aptamer fragment 1 and aptam...
Embodiment 2
[0029]1) Mix aptamer fragment 1, dye-labeled aptamer fragment 2 and water-soluble conjugated polymer at a molar ratio of 1:1:10 to form an aptamer / polymer complex, and then add it to the buffer solution In this method, the fluorescence detection is carried out with 404 nm as the excitation wavelength, the fluorescence emission band of the complex is recorded, and the ratio I of the fluorescence intensity at 525 nm to 440 nm is calculated 525 / I 440 ; The aptamer fragment 1 and the aptamer fragment 2 are specific single nucleotide sequences, and the 3' end of the aptamer fragment 2 is labeled with fluorescein, and the labeling method is: 3'-6-FAM labeling.
[0030] 2) Add aptamer fragment 1 and aptamer fragment 2 and the test sample containing thrombin into the binding solution and place it at 37°C for 40 minutes to form a thrombin / aptamer complex.
[0031] 3) Adding a water-soluble conjugated polymer to the above complex, thrombin induces and combines with aptamer fragment 1 ...
Embodiment 3
[0032] Example 3 Specific Analysis of Thrombin Detection
[0033] Add 2.5 μL of aptamer fragment 1 at a concentration of 10 μM and 2.5 μL of aptamer fragment 2 at a concentration of 10 μM to each centrifuge tube, then add 1 μL of thrombin (Thro) at a concentration of 10 μM, 6.8 μL Bovine serum albumin (BSA) at a concentration of 0.1 mg / ml, 2 μL of lysozyme (Lys) at a concentration of 5 μM, and 1.6 μL of immunoglobulin (IgG) at a concentration of 1 mg / ml, so that the final concentration of each protein was 20 nM, The final concentration of aptamer fragment 1 and aptamer fragment 2 is 50 nM; finally add 2.5 μL of a water-soluble conjugated polymer with a concentration of 50 μM, and perform fluorescence detection under the environment of pH=10.6, 0.1 M CBS buffer solution , see the result image 3 , it can be seen from the figure that the FRET of the blank and the addition of non-specific protein are similar and relatively high, and the FRET of the addition of thrombin is signif...
PUM
Login to view more Abstract
Description
Claims
Application Information
Login to view more - R&D Engineer
- R&D Manager
- IP Professional
- Industry Leading Data Capabilities
- Powerful AI technology
- Patent DNA Extraction
Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic.
© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap