QCM detection method for detecting lysozyme based on multiple signal amplification technologies and application

A detection method and technology for technical detection, applied in biochemical equipment and methods, microbial determination/inspection, etc., can solve the problems of accurate quantitative analysis, low number of tumor markers, etc., and achieve the effect of simple operation and low cost

Inactive Publication Date: 2017-12-01
QINGDAO UNIV
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Problems solved by technology

There are more than 100 tumor markers that have been discovered, but the number of tumor markers in the early stages o

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  • QCM detection method for detecting lysozyme based on multiple signal amplification technologies and application
  • QCM detection method for detecting lysozyme based on multiple signal amplification technologies and application

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Embodiment 1

[0073] Ultrasensitive detection of lysozyme based on multiple signal amplification techniques

[0074] 1. Modification of magnetic bead carrier

[0075] (1) Activated magnetic beads

[0076] The magnetic beads were washed three times with imidazole hydrochloride buffer (0.1 M, pH 6.8), added to 100 μL of imidazole buffer containing 0.2 mol / LEDC, and shaken at 37° C. for 2 h.

[0077] (2) Magnetic bead carrier modification

[0078] Add 15 μL to the activated magnetic beads with a concentration of 1.0×10 -7 mol / L lysozyme aptamer DNA, shake at 37°C for 16h. The DNA not attached to the magnetic beads was removed by magnetic separation, washed with Tris-HCl, added to the main DNA solution, and incubated for 1 h. Magnetic separation, washing with Tris-HCl, and then adding 30 μL of different concentrations (1.0×10 -15 mol / L, 1.0×10 -14 mol / L, 1.0×10 -13 mol / L, 1.0×10 -12 mol / L, 1.0×10 -11 mol / L) of lysozyme, incubated at 37°C for 1 h, magnetically separated, and the superna...

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Abstract

The invention discloses a QCM detection method for detecting lysozyme based on multiple signal amplification technologies and application. The QCM detection method comprises the following steps that DNA hybridizes with lysozyme aptamer partially in a complementary mode, and through the specific binding reaction of the lysozyme and the lysozyme aptamer, the DNA is released; a Y-shaped structure is formed by complementary hybrid of the released DNA with hairpin DNA and assistant DNA modified on a gold leaf, and under the action of restriction enzyme, the hairpin DNA is cut and opened through specific identification sites; under the action of DNA ligase and DNA polymerase, using locking-ring-shaped DNA as a template chain, polymerization growing along the opened hairpin DNA is carried out, and a single chain with a large number of repeated sequences is formed; a signal probe marked with biotin hybridizes with the generated repeated sequences in a complementary mode, and after binding with streptavidin marked by HRP, hydrogen peroxide is catalyzed to oxidize 4-chloro naphthol, and precipitation reaction is generated; and accordingly the chip surface quality is increased, and the high-sensitivity detection of the QCM to the lysozyme is realized.

Description

technical field [0001] The invention relates to a QCM detection method and application for detecting lysozyme based on multiple signal amplification technologies, belonging to the technical field of biomolecular detection. Background technique [0002] Tumor is one of the malignant diseases that seriously threaten human life and health. At present, clinical diagnosis methods mainly rely on imaging to detect the morphology of tumor cells, with low resolution. Tumor markers are an important indicator of tumor diagnosis, and their detection is of great significance to the early diagnosis and treatment of diseases, and to the prognosis. There are more than 100 tumor markers that have been discovered, but the number of tumor markers in the early stages of cancer patients is generally very small, and it is difficult to accurately quantitatively analyze them with conventional detection methods. Therefore, it is of great scientific significance and clinical value for the early dia...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12Q1/34
CPCC12Q1/682C12Q1/6825C12Q2521/301C12Q2525/301C12Q2531/125C12Q2563/131
Inventor 王宗花孙文博夏建飞张菲菲
Owner QINGDAO UNIV
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