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Detection kit for detecting biomarker of bladder cancer

A technology of biomarkers and diagnostic kits, applied in the fields of biotechnology and medicine, can solve problems such as unsatisfactory sensitivity and specificity, and achieve the effect of avoiding pain and high sensitivity

Inactive Publication Date: 2020-05-15
北京恩泽康泰生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In addition to urine cytology, the current diagnosis and detection of bladder cancer recurrence through urine examination mainly include urinary nuclear matrix protein 22, bladder cancer tumor antigen, immune-cytological examination, etc., but the sensitivity and specificity are not ideal

Method used

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  • Detection kit for detecting biomarker of bladder cancer
  • Detection kit for detecting biomarker of bladder cancer
  • Detection kit for detecting biomarker of bladder cancer

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0047] Example 1 Separation method of urine exfoliated cells

[0048] Transfer 40ml of human morning urine sample to a 50ml sterile centrifuge tube, centrifuge at room temperature at 2000g for 20min, carefully take out the centrifuge tube, pour off the supernatant slowly, and be careful not to pour off the exfoliated cells in the lower layer. Add 15ml of 1×PBS, invert the centrifuge tube 3-4 times to mix the sample, centrifuge at 2000g room temperature for 20min, carefully take out the centrifuge tube, slowly pour off the supernatant, be careful not to pour off the exfoliated cells in the lower layer, add about 100μl of 1 Resuspend in ×PBS and transfer to a 1.5ml EP tube.

Embodiment 2

[0049] Example 2 Screening of differentially expressed genes

[0050] In order to screen biomarkers related to the diagnosis of bladder cancer, the transcriptome data published in TCGA (https: / / cancergenome.nih.gov / ) and pubmed of NCBI (https: / / www.ncbi.nlm. The candidate markers obtained by consulting relevant literature in nih.gov / pubmed / ) are as follows: CA9, CRH, HOXA13, IGF2, RPS21, CDK1, UBE2C. In addition, at least one of the three internal reference genes SLC25A6, ACTB and GAPDH was selected as an internal reference to normalize the above seven genes.

Embodiment 3

[0051] Example 3 Verify the primers of selected gene markers by real-time fluorescent PCR

[0052] 1. Design and synthesis of primers and probes

[0053] Combining Primer Premier 5 software and Primer-BLAST (NCBI), primers and probes for 7 bladder cancer markers including CA9, CRH, HOXA13, IGF2, RPS21, CDK1 and UBE2C and 3 reference genes including SLC25A6, ACTB and GAPDH were designed . The design principles are as follows: 1) The amplified fragment is less than 150bp; 2) The amplified fragment spans at least one exon; 3) The Tm value of the probe is at least 5°C higher than that of the primer. The designed primers and probe sequences were synthesized by the company, in which the 5' end of the probe was labeled with FAM group, and the 3' end was labeled with BHQ1.

[0054] The nucleotide sequence of the designed primer probe is the sequence shown in SEQ ID NO: 1-30.

[0055] 2. Reverse transcription and qPCR detection

[0056] Takara's PrimeScriptTMRT reagent Kit (Perfect...

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Abstract

The invention provides a detection kit for detecting a biomarker of the bladder cancer. The detection kit includes a primer and probe for detecting the biomarker of the bladder cancer, the biomarker of the bladder cancer is at least one selected from genes of CA9, IGF2, CDK1, UBE2C, CRH, RPS21 and HOXA13. Furthermore, the detection kit further includes a primer and probe for detecting a referencegene, the reference gene is selected from SLC25A6, ACTB or GAPDH. The invention also provides a bladder cancer-diagnosing diagnostic kit including the detection kit. The diagnostic kit is capable of diagnosing whether an individual has the bladder cancer or not and the possibility of recurrence accurately, and has high sensitivity, and the functions of auxiliary diagnosis and recurrence monitor ofnon-invasive bladder cancer are achieved.

Description

technical field [0001] The invention belongs to the field of biotechnology and medicine, in particular, relates to biomarkers and kits for bladder cancer diagnosis and recurrence monitoring, more specifically, relates to detection kits for detecting bladder cancer biomarkers and for diagnosis Diagnostic kit for bladder cancer. Background technique [0002] Bladder cancer (Bladder Cancer, BC) refers to malignant tumors that occur on the bladder mucosa. It is the most common tumor in the genitourinary system in my country. The incidence is increasing year by year and tends to be younger. Bladder cancer can be divided into muscle invasive bladder cancer (MIBC) and muscle noninvasive bladder cancer (NMIBC) according to the depth of disease invasion. MIBC has a higher risk of disease progression, while NMIBC is more likely to relapse. [0003] Cystoscopy and cytology are still the main techniques for the diagnosis of bladder cancer, but the former is invasive and expensive, whil...

Claims

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Application Information

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IPC IPC(8): C12Q1/6886C12N15/11
CPCC12Q1/6886C12Q2600/166
Inventor 秦宏亮陈亚庆李鸿艳秦伟伟刘翔陈苏红孔关义赵立波
Owner 北京恩泽康泰生物科技有限公司
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