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Detection kit for prostate specific antigen, detection method and application thereof

A prostate-specific, detection kit technology, which can be used in biological tests, measuring devices, material inspection products, etc., and can solve problems such as cumbersome steps and limitations

Active Publication Date: 2019-09-06
LINYI UNIVERSITY
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  • Abstract
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  • Claims
  • Application Information

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Problems solved by technology

DNA-assisted protein detection technology can improve the performance of detection technology, such as polymerase chain reaction, rolling circle amplification, etc., but the cumbersome steps and possible false positives of polymerase amplification limit its application.

Method used

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  • Detection kit for prostate specific antigen, detection method and application thereof
  • Detection kit for prostate specific antigen, detection method and application thereof
  • Detection kit for prostate specific antigen, detection method and application thereof

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Embodiment 1

[0050] Equipment: high-speed refrigerated centrifuge (16R, Zhuhai Heima Medical Instrument Co., Ltd.), ultrapure water machine (SybergyUV, Merck Millipore), ultraviolet-visible spectrophotometer (Cary60, Agilent Technologies, USA), fluorescence spectrophotometer ( F-4600, Japan Hitachi High-Tech Corporation), agarose horizontal electrophoresis tank (DYCP-31DN, Beijing Liuyi).

[0051] Reagents used: PSA antibody, SMCC (succinimide-4-(N-maleimide)cyclohexane-1-1 hydroxyester), TCEP (tris(2-carboxyethyl)phosphine), mercapto Modified DNA1, thiol-modified DNA2, MB (DNA is biosynthesized by Bao, MB contains an rA base that can be recognized by ionase and is modified with FAM and BHQ groups).

[0052] 1. Antibody modified DNA

[0053] Antibody and SMCC (suitable molar ratio 1:10-20) were reacted in PBS buffer at room temperature for 2 hours, then purified with filter membrane (molecular weight cut-off 10,000MW), washed three times with PBS buffer; Add 2 μL TCEP (100 mM) to sulfhyd...

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Abstract

The invention provides a detection kit for a prostate specific antigen, a detection method and an application thereof, which relate to the technical field of antigen detection. The kit and the detection method of the invention utilize a secondary antibody to modify DNA, form a Y-shaped antigen-antibody-DNA hybridization chain by means of proximity ligation with the antigen, and converts the antigen input into DNA signal output by using the antigen-antibody-DNA hybridization chain as a signal converter; meanwhile, a chain substitution reaction is combined with a hybridization chain type reaction, and two-stage amplification of the signal is achieved; and every two adjacent two-end sequences (hairpin DNA H3 and hairpin DNA H4) on long-chain DNA are assembled to form a magnesium ion DNA enzyme, the magnesium ion DNA enzyme cuts molecular beacons in a solution in the presence of magnesium ions, the cut-off molecular beacons are released and fluoresce, new molecular beacons are combined with the magnesium DNA enzyme and cut off by the same, and a fluorescent signal is cyclically amplified to realize the third round of signal amplification. The detection kit and the detection method greatly improve the detection efficiency and sensitivity, so that a detection limit is up to 0.73 pg mL<-1>.

Description

technical field [0001] The invention belongs to the technical field of antigen detection, and in particular relates to a detection kit, detection method and application of prostate specific antigen. Background technique [0002] Antigen detection has important reference value in clinical diagnosis. Surface molecules of human and animal cells: including various differentiation antigens (such as CD antigens), allotype antigens (blood group antigens or MHC antigens), virus-associated antigens and tumor-associated temperament antigens on the cell surface. The detection of these antigens is of great significance to the classification of various cells, the study of differentiation process and function, the diagnosis and pathogenesis of various immune-related diseases. [0003] Currently, antigen detection methods mainly include traditional enzyme-linked immunoassay (ELISA), radioimmunoassay (RIA), fluorescence immunoassay (FIA), chemiluminescence immunoassay (CLIA), and automated...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/577G01N33/53
CPCG01N33/577G01N33/5308G01N2800/342
Inventor 张宁波井文倩刘静周宏
Owner LINYI UNIVERSITY
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