Quantitative detection kit and method for exon mutation of epidermal growth factor receptor (EGFR) relevant to lung cancers

An epidermal growth factor, quantitative detection technology, applied in the direction of microbial determination/inspection, biochemical equipment and methods, etc., can solve the problems of deterioration of the disease, delaying the timing of treatment, etc., to reduce the false positive or false negative rate and save treatment time. , The effect of reducing the requirements for sample collection

Active Publication Date: 2011-08-03
北京迪安医学检验实验室有限公司
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Problems solved by technology

Blind drug use without target detection or false-negative or false-positive test results may not only lead to

Method used

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  • Quantitative detection kit and method for exon mutation of epidermal growth factor receptor (EGFR) relevant to lung cancers
  • Quantitative detection kit and method for exon mutation of epidermal growth factor receptor (EGFR) relevant to lung cancers
  • Quantitative detection kit and method for exon mutation of epidermal growth factor receptor (EGFR) relevant to lung cancers

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Experimental program
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Effect test

Embodiment 1

[0042] Example 1: Reagents.

[0043] (1) DNA extraction reagents:

[0044] Purchased from QIAGEN Company.

[0045] (2) Reaction solution:

[0046] PCR Buffer: purchased from Fermentas, USA;

[0047] Primers SEQ ID NO: 1-12, synthesized by Shanghai Yingjun Biotechnology Co., Ltd.;

[0048] MgCl 2 : purchased from the U.S. Fermentas company;

[0049] 0.2mM dNTPs: purchased from Fermentas, USA;

[0050] 2U / μL Taq DNA polymerase: purchased from Fermentas, USA.

[0051] (3) Reagent for single-strand purification:

[0052] 75% (v / v) ethanol solution: purchased from Hangzhou Changzheng Chemical Reagent Co., Ltd.;

[0053] 0.2M NaOH: purchased from Shanghai Shisi Hewei Chemical Co., Ltd.;

[0054] 10mM Tris-Acetate (pH 7.6): Tris-base was purchased from Sigma, USA, and anhydrous acetic acid was purchased from Hangzhou Chemical Reagent Co., Ltd.;

[0055] Binding buffer: 10mM Tris-HCl (Tris-base was purchased from Sigma, USA; hydrochloric acid was purchased from Hangzhou Chem...

Embodiment 2

[0062] Embodiment 2: detection method.

[0063] Instruments: Bio-Rad S1000 PCR instrument, Beckman Microfuge 22R desktop micro-centrifuge, Beijing Liuyi agarose gel electrophoresis instrument, Shanghai Peiqing gel imaging system, QIAGEN PyroMark Q96ID sequencer. (1) Extract tissue DNA from paraffin specimens, the specific steps are as follows: place the paraffin specimens in xylene to remove paraffin; add lysis buffer and proteinase K, digest the tissue under denaturing conditions, and lyse the cells; incubate at 90°C, reverse Formalin cross-linking; pass the lysate through the silica gel membrane to adsorb DNA to the silica gel membrane, add rinsing solution to wash the impurities, and finally elute the high-purity and concentrated DNA from the silica gel membrane to obtain the genomic DNA collection solution.

[0064] (2) using the DNA obtained in step (1) as a template, and using EGFR-specific primers to perform PCR amplification;

[0065] The method for amplifying the EGF...

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Abstract

The invention discloses a quantitative detection kit for exon mutation of epidermal growth factor receptor (EGFR) relevant to lung cancers, which comprises a primer for amplifying exons of EGFR genes 18, 19, 20, 21, a sequencing primer and a magnetic bead marked by avidin. The invention also discloses a preparation method and a using method of the kit. The detection method in the invention not only has high sensitivity so that 1% of mutation can be detected, but also has visual result and simple, accurate and rapid interpretation, greatly reduces the false negative rate of the detection result and avoids invalid targeted medicine selections of clinical patients, thereby saving valuable treating time for the patients and improving the life quality of the patients.

Description

technical field [0001] The invention belongs to the field of molecular pathological diagnosis, and specifically relates to a method for accurately and quantitatively detecting the mutation of epidermal growth factor receptor EGFR related to lung cancer. Background technique [0002] Lung cancer is very harmful and has a high mortality rate. It is currently the number one cause of cancer death in the world. In 1995, 600,000 people died of lung cancer in the world, and the number is increasing every year. In 2003, the death rate announced by the World Health Organization (WHO) The incidence rate is 1.1 million per year, and the incidence rate is 1.2 million per year. 75% to 80% of lung cancers are non-small cell lung cancers. Treatment methods such as radiotherapy and chemotherapy for patients with advanced lung cancer not only reduce the quality of life, but also have poor curative effect. Iressa (Gefitinib) is currently the most successful clinically used drug for targeted ...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
Inventor 任绪义张峰虞闰六
Owner 北京迪安医学检验实验室有限公司
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