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Molecular reagent for screening bladder cancer by using urine

A bladder cancer and urine technology, applied in the field of epigenetic research, can solve the problems of subjective interpretation by the naked eye, subjective judgment, complicated operation, etc., and achieve accurate and reliable test results, high clinical positive predictive value and negative predictive value. effect of value

Inactive Publication Date: 2018-06-08
上海纽思格生物医学科技有限公司
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Problems solved by technology

The above-mentioned laboratory diagnostic products all have shortcomings such as low sensitivity, low specificity, cumbersome operation, and subjective judgment.
[0004] The existing bladder cancer molecular diagnosis product "The UroVysion Bladder Cancer Kit" is based on in situ hybridization microscopy. The experimental process is complicated and cannot be standardized. The interpretation is subjective and requires professional operation and judgment.

Method used

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  • Molecular reagent for screening bladder cancer by using urine
  • Molecular reagent for screening bladder cancer by using urine

Examples

Experimental program
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Effect test

Embodiment example 1

[0029]Implementation case 1: 50 samples of urine (not less than 10ml) from inpatients in the Urology Department of Shanghai Tenth People’s Hospital, including 25 bladder cancer patients, 11 prostate patients, and the rest were non-tumor patients of the urinary system such as benign prostatic hyperplasia, Prostatitis, urethritis and other patients. Through the technical scheme of the present invention (the above-mentioned classical two-step purification method is used for nucleic acid extraction and bisulfite conversion), the following primer-probe combinations are used for detection: VIM (FAM fluorescence), TMEFF2 (VIC fluorescence), GDP15 (ROX fluorescence) , HSPA2 (CY5 fluorescence), ABL1 (NED fluorescence), all use MGB probes with high hybridization resolution, run the program on ABI7500: 50°C for 2min, 95°C for 5min, 95°C for 10sec, 60°C for 40sec, cycle 40-45 Second, 5-color fluorescence detection. Of course, the same effect can be obtained by dividing the single-color f...

Embodiment example 2

[0046] Example 2: 38 samples of urine (not less than 10ml) were collected from the Urology Department of Shanghai Pudong Hospital, including 9 bladder cancer patients, 20 prostate patients, and 9 non-tumor patients in the urinary system. Through the technical scheme of the present invention (the above-mentioned one-step method is used for nucleic acid extraction and bisulfite conversion), the combination of primers and probes in Example 1 is used for detection, but the probes are all labeled with FAM: each nucleic acid is divided into tubes for detection, GDF15 and VIM The result is as figure 2 : All 9 bladder cancer patients were detected, 1 of 20 prostate patients was detected positive, and 9 non-tumor patients had no detection signal. The positive predictive value was 90%, and the negative predictive value was 100%.

[0047] HSf: 5'-ttt cgt ttt aac gtcgtt cgt tt-3'

[0048] HSr: 5'-ccg acg cac gaa taa ata ata cc-3'

[0049] HSp: 5’-tcg gtt atc ggt atc gat- 3’

[0050] ...

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Abstract

The invention belongs to the field of biotechnology molecular diagnosis, and provides a molecular diagnosis method of noninvasive detection of bladder cancer by using urine. The method comprises the following steps of 1, urine nucleic acid extraction and purification; 2, DNA hydrosulphite conversion reaction and subsequent purification; 3, methylation specific real-time fluorescence PCR detectionon a plurality of gene promotor regions; 4, multi-gene result analysis. The steps 1 and 2 can be merged into one step. The multi-gene comprises the promotor regions of the following genes including VIM, TMEFF2, HSPA2, GDF15, ABL1, CRH, IGF2, ANXA10, UPK1B, ACTB and the like; the gene combination detection results can be used for bladder cancer diagnosis and bladder cancer recurrence rate prediction.

Description

technical field [0001] The present invention belongs to the field of biomedicine, especially the field of molecular diagnosis related to non-invasive tumor screening. Development-related, including detection, prediction, prognosis, drug efficacy, lymph node metastasis, etc. Background technique [0002] Bladder cancer is the most common malignant tumor of the urinary system, and its incidence rate ranks first among malignant tumors of the urinary system. It is generally believed that it is related to frequent exposure to carcinogens such as naphthylamine and benzidine. Common dyes, rubber, plastic products, paints, detergents, etc. in daily life have potential carcinogenic risks. for environmental tumors. In addition, smoking is also an independent risk factor for bladder cancer. Bladder cancer occurs more frequently in people over the age of 50, and more men than women. At present, the curative effect of surgery, radiotherapy and chemotherapy is not satisfactory, and the...

Claims

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Application Information

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IPC IPC(8): C12Q1/6886C12N15/11C12N15/10
CPCC12Q1/6886C12N15/1013C12Q2600/118C12Q2600/154
Inventor 周科隆宋军
Owner 上海纽思格生物医学科技有限公司
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