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Method for detecting tiny amount of prostate cancer specific membrane antigen in circulating blood, and kit thereof

A prostate cancer, specific technology, applied in the field of protein detection, can solve the problems of high false positive rate and high environmental purity requirements of antibody specificity

Inactive Publication Date: 2015-08-19
SUZHOU YOULIN BIO TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The first problem to be solved by the present invention is to overcome the problems that the specific detection of human prostate cancer-specific membrane antigen requires high environmental purity, very high antibody specificity and high false positive rate, and provides a circulating blood Prostate cancer-specific membrane antigen detection method, the method is easy to use, sensitive, specific, good repeatability, quick results, no need for complicated instruments and special skills, and can be directly detected by using the sample to be tested without precision purification , there will be no mutual interference between the aptamer and the target molecule (prostate cancer-specific membrane antigen) and other non-specific proteins

Method used

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  • Method for detecting tiny amount of prostate cancer specific membrane antigen in circulating blood, and kit thereof
  • Method for detecting tiny amount of prostate cancer specific membrane antigen in circulating blood, and kit thereof
  • Method for detecting tiny amount of prostate cancer specific membrane antigen in circulating blood, and kit thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0068] Example 1 Preparation of prostate cancer cell protein samples and their controls for detection

[0069] Cultivate LNCaP (PSMA+) cells in vitro, digest the adherent cells with trypsin to suspend the cells, centrifuge to remove the culture medium, then add normal saline to prepare a single cell suspension, and take a certain amount of cells (10 7 )conduct experiment. According to the manufacturer's instructions (Abio), the total protein of LNCaP cells was extracted with the total cell protein extraction reagent, and the amount of quantitative total protein was measured. Then, the total protein was diluted with PBS buffer (pH7.2), and the equal volumes of solutions of each dilution contained 5 (Group A), 10 4 (Group B), 10 3 (Group C), 10 2 (Group D) and 10 1 (Panel E) Total protein of LNCaP prostate cancer cells.

[0070] According to the above method, the total protein of PC3 (PSMA-) cells was extracted, and after dilution, equal volumes of solutions of each diluti...

Embodiment 2

[0071] Example 2 Comparison with the method of detecting prostate cancer specific membrane antigen by ELISA

[0072] According to the determined conditions, the LNCaP cell total protein (A, B, C, D, E group dilutions) prepared in Example 1 known to contain the target molecule (PSMA protein) was detected by ELISA respectively, and according to the intensity of the color reaction Weak to determine the presence of target protein molecules. Target molecule (PSMA) expression negative PC3 cell total protein (A 0 , B 0 、C 0 、D 0 ,E 0 Group diluent) is the corresponding control group. The results are as follows Figure 1.1 As shown, the OD value of total protein in PC3 (PSMA-) cells with negative target molecule expression is very low, which can be used as a background control, and its detection line is used as a standard baseline for detection (higher than this line is a positive test result); while in In the triplicate detection of the serially diluted protein samples of LNCaP...

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Abstract

The invention provides a method for detecting a tiny amount of a prostate cancer specific membrane antigen in circulating blood. The method comprises the following steps: respectively mixing a magnetic particle coupled specific PSMA (prostate specific membrane antigen) aptamer and a nanogold particle coupled specific PSMA antibody with a sample to be detected, detecting, and identifying and / or quantifying a target molecule (PSMA). The invention also relates to a pretreatment agent of the blood sample to be detected, and a detection kit which are used for the method. The method has the advantages of simple use, sensitivity, strong specificity, good repeatability, fast result obtaining, and no complex apparatuses or special skills, and still keeps practical sensitivity and specificity when the sample to be detected is directly detected without precise purification, so the detection operation is greatly convenient, and practical industrial application of the method is popularized.

Description

technical field [0001] The invention relates to the technical field of protein detection methods, in particular, the invention relates to a specific detection method for detecting, identifying and / or quantifying a small amount of prostate cancer-specific membrane antigen in human circulating blood. In addition, the present invention also relates to kits used in the method and applications thereof. Background technique [0002] Prostate cancer is one of the most common fatal cancers in elderly men. Surgical resection is the first choice in the comprehensive treatment of prostate cancer, but the recurrence rate and metastasis rate after resection of prostate cancer are high: cancer cells are easy to enter the peripheral blood from the primary tumor Tumor metastasis occurs, which is the main cause of treatment failure in prostate cancer. A large number of experiments have confirmed that the detection of Circulating Tumor Cells (CTCs) in circulating blood will be helpful for th...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/574G01N33/566
CPCG01N33/57434G01N33/54326G01N33/54346G01N33/566
Inventor 游绍进马杰
Owner SUZHOU YOULIN BIO TECH
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