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Method for portably detecting vascular endothelial growth factor by using hybrid chain reaction, and nucleic acid sequence used by method

A portable detection and hybridization chain reaction technology, applied in the field of nucleic acid sequence, can solve the problems that are difficult to apply to ordinary people's daily detection and cumbersome operations

Inactive Publication Date: 2016-11-09
FUJIAN AGRI & FORESTRY UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although this series of aptasensors can successfully detect VEGF in the laboratory, the storage of the enzyme requires strict temperature control, cumbersome operations during the use of the enzyme, and the need for large and expensive detection instruments and professional expertise. Operators, so the application of these methods is often limited to the laboratory, it is difficult to apply to the daily detection of ordinary people

Method used

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  • Method for portably detecting vascular endothelial growth factor by using hybrid chain reaction, and nucleic acid sequence used by method
  • Method for portably detecting vascular endothelial growth factor by using hybrid chain reaction, and nucleic acid sequence used by method
  • Method for portably detecting vascular endothelial growth factor by using hybrid chain reaction, and nucleic acid sequence used by method

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0058] Embodiment one: measure when the concentration of VEGF standard solution is 50pg / mL, corresponding blood glucose meter reading—a

[0059] Step 1): Add 10 μg / mL of VEGF antibody solution dropwise onto a polystyrene microwell plate at an amount of 50 uL / well, incubate at 37.0°C for 1 hour, and rinse the polystyrene microwell plate 3 times with distilled water , each time for 3 minutes, then add 1%-5% bovine serum albumin to the polystyrene microwell plate at an amount of 50uL / well, shake at 37.0°C for 1 hour, and then rinse the polystyrene with distilled water again Microplate 3 times, 3 minutes each.

[0060] Step 2): VEGF is dissolved in a PBS buffer solution with a concentration of 0.1M and a pH of 7.5, and is prepared into a VEGF standard solution with a concentration of 50pg / mL, and the VEGF standard solution is added to the step in an amount of 50 μL / well 1) In the obtained polystyrene microwell plate, after incubating at 37.0° C. for 30 minutes, the polystyrene mi...

Embodiment 2

[0080] Embodiment two: measure when the concentration of VEGF standard solution is 50pg / mL, corresponding blood glucose meter reading—b

[0081] Step 1): Add 20 μg / mL of VEGF antibody solution dropwise to a polystyrene microwell plate at an amount of 30 uL / well, incubate at 37.5°C for 0.5 hours, and rinse the polystyrene microwell plate twice with distilled water , each time for 5 minutes, then add 1%-5% bovine serum albumin to the polystyrene microwell plate with an addition amount of 30uL / well, shake at 37.5°C for 1.5 hours, and then rinse the polystyrene with distilled water again Microplate 2 times, 5 minutes each time.

[0082] Step 2): VEGF is dissolved in a PBS buffer solution with a concentration of 0.2M and a pH of 7.0, and is prepared into a VEGF standard solution with a concentration of 50pg / mL, and the VEGF standard solution is added to the step in an amount of 30 μL / well 1) In the obtained polystyrene microwell plate, after incubating at 37.5° C. for 60 minutes, ...

Embodiment 3

[0102] Example 3 Determination When the concentration of the VEGF standard solution is 50pg / mL, the corresponding blood glucose meter reading—c

[0103] Step 1): Add 15 μg / mL of VEGF antibody solution dropwise to a polystyrene microwell plate at an amount of 40 uL / well, incubate at 37.3°C for 3 hours, and rinse the polystyrene microwell plate 4 times with distilled water , each time for 4 minutes, then add 1%-5% bovine serum albumin to the polystyrene microwell plate with the addition amount of 40uL / well, shake at 37.3°C for 2 hours, after that, wash the polystyrene with distilled water again Microplate 4 times, 4 minutes each.

[0104] Step 2): VEGF is dissolved in a PBS buffer solution with a concentration of 0.05M and a pH of 7.6, and is prepared into a VEGF standard solution with a concentration of 50pg / mL, and the VEGF standard solution is added to the step in an amount of 100 μL / well. 1) In the obtained polystyrene microwell plate, after incubating at 37.3° C. for 50 mi...

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Abstract

The invention relates to a method for portably detecting a vascular endothelial growth factor by using a hybrid chain reaction, and a nucleic acid sequence used by the method. The method for detecting the vascular endothelial growth factor comprises the following steps: VEGF, an aptamer and an antibody form an antibody-protein-aptamer sandwich structure on polyethylene micro-pores, the tail end of the aptamer includes a 18-base extended sequence, two fragments of synthesized sucrose transferase labeled auxiliary probes initial a chain hybrid amplification reaction with the 18-base extended sequence as a template in order to immobilize a large amount of sucrose transferase in the micro-pores, the sucrose transferase catalyzes conversion of sucrose into glucose, and the concentration of the generated glucose is indicated by a glucometer. The detection method has the advantages of portability, simplicity, convenience in operation, high sensitivity, high selectivity, and realization of effective detection of the content of the vascular endothelial growth factor in a target to be analyzed.

Description

technical field [0001] The invention relates to the field of detection and analysis, in particular to a method for portable detection of vascular endothelial growth factor by hybridization chain reaction and the nucleic acid sequence used in the method. Background technique [0002] Vascular endothelial growth factor (VEGF) is a new type of signaling protein found in cells. It can increase the permeability of microvessels and venules, and is an important regulatory factor for promoting angiogenesis. The overexpression of VEGF is also closely related to many diseases, such as lung cancer, breast cancer, rectal cancer, psoriasis, proliferative retinopathy and rheumatoid arthritis, etc. Therefore, VEGF is often used as an important biomarker in the diagnosis of clinical diseases or as an indicator of prognosis. [0003] At present, conventional VEGF detection methods are mainly based on immunoassay, including immunohistochemistry, radioimmunoassay and so on. This type of meth...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/74C12N15/115
CPCC12N15/115C12N2310/16G01N33/74
Inventor 朱希许惠凤杨桂娣寇芳霞
Owner FUJIAN AGRI & FORESTRY UNIV
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