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Novel prostate-specific antigen detection kit

A prostate-specific detection kit technology, applied in the field of immunoanalysis medicine, can solve the problems of limited detection objects, inability to meet the sensitive detection of low-abundance proteins in serum, unfavorable early tumor disease, and extensive screening

Inactive Publication Date: 2015-11-11
LINYI UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, although DNA molecular machines based on isothermal amplification technology are used for the detection of tumor diseases due to the excellent designability and obvious amplification effect of DNA, the current detection objects are limited to nucleic acid tumor markers or tumor cells themselves.
Nucleic acid tumor markers basically exist only inside tumor cells, which means that operations such as tissue biopsy and cell extraction are required in the process of disease detection. The detection process is relatively traumatic to patients, which is not conducive to early and extensive screening of tumor diseases
However, due to the limitations of detection methods, the detection of serum tumor markers with relatively small trauma is far lower than the sensitivity of nucleic acid tumor markers, which cannot meet the sensitive detection of low-abundance proteins in serum.

Method used

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Examples

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Embodiment 1

[0024] The following are specific examples for implementing the present invention, which serve to further clarify the content of the present invention and make it easier for readers to understand, but do not constitute a limitation or limitation on the scope of protection claimed by the present invention.

[0025] Experimental conditions:

[0026] Fluorescence spectrophotometer (F-4500 type, Japan, Hitachi company), the deoxyribonucleotide sequence used and the mixture of four deoxynucleotide triphosphates (dNTP) (Dalian Bao Biological Engineering Co., Ltd.), prostate specific antigen And antibody (Shanghai Lingchao Biological Co., Ltd.), biotin-NHS, bovine serum albumin (Sigma Reagent Company, USA), cutting enzyme Nb.BtSI, polymerase Klenow fragment, NEBuffer buffer (New England Biotechnology Co., Ltd.) , sodium carbonate, sodium bicarbonate and other basic drugs (Sinopharm Chemical Reagent Co., Ltd.), 10×Klenow buffer composition: 100mM Tris-HCl, 70mM MgCl 2 And 1mM DTT, pH...

Embodiment 2

[0038] Example 2 The using method of kit of the present invention:

[0039] 1. Take out the kit from the refrigerator at 4°C, equilibrate at room temperature for 20 minutes, take a bottle of concentrated lotion, add distilled water (1mL+19mL) for later use.

[0040] 2. Take the microplate out of the sealed bag, fill the wells with the washing solution, let it stand for 10-20 seconds, shake off the washing solution, repeat the plate washing, and finally pat dry on clean absorbent paper.

[0041] 3. Set up 2 wells for each series of standard substance concentrations, and 2 wells for each sample well. Take 50 μL each of the standard substance or sample solution in the corresponding plate wells, fully oscillate and mix with a micro-oscillator, and cover the reaction plate with a self-adhesive film. The reaction plate was then incubated at 37°C for 30 minutes.

[0042] 4. Shake off the reaction solution carefully, then wash five times as in step 2.

[0043] 5. Take 100 μL of clea...

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Abstract

The invention relates to the medical field of immunoassay, and specifically, the invention innovatively designs a prostate-specific antigen detection kit based on second antibody modified shear enzyme initiated DNA molecular machine cycle amplification. The kit provided by the invention includes: an anti-PSA antibody coated solid phase carrier, a shear enzyme labeled anti-fPSA antibody, a reaction solution (DNA molecular machine) on which the shear enzyme acts, and a negative control solution and a positive control solution. The kit provided by the invention has high sensitivity, and compared with the existing optical immunoassay detection method, can improve the analysis sensitivity and reproducibility, and reduce the antibody dosage.

Description

technical field [0001] The invention relates to the field of immunoanalysis medicine, in particular, the invention is a reagent kit for immunoassay and detection of prostate specific antigen based on DNA molecular machine cycle amplification technology triggered by secondary antibody modified shear enzyme. Background technique [0002] Prostate cancer (CaP) is currently the most common cancer in men, and if it can be diagnosed at an early stage, it can be effectively dealt with and treated. Prostate-specific antigen (PSA) has been used as the main protein biomarker for clinical diagnosis of prostate disease since it was developed decades ago. PSA mainly exists in free PSA and bound state PSA in serum, the free state is called fPSA, the bound state is called (tPSA)PSA, and the level is 4ngmL -1 or lower is considered normal, and more than 4ngmL -1 considered to be dangerous. [0003] Traditional methods for detecting prostate specific antigen include enzyme-linked immunoas...

Claims

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Application Information

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IPC IPC(8): G01N33/68G01N33/574G01N33/543G01N33/535
CPCG01N33/6893G01N33/535G01N33/543G01N33/57434
Inventor 周宏张宁波刘静张书圣
Owner LINYI UNIVERSITY
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