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LAMP primer group CK19-115 for quickly detecting lymphatic metastasis of head and neck squamous cell carcinoma

A technique for squamous cell carcinoma and lymph node metastasis, which is applied in the fields of life science and clinical medicine, and can solve problems such as unfavorable intraoperative procedures.

Active Publication Date: 2017-05-24
BEIJING CHILDRENS HOSPITAL AFFILIATED TO CAPITAL MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, conventional qRT-PCR analysis generally takes 3-4 hours, which is not conducive to intraoperative implementation. Therefore, a molecular biological method that can accurately and quickly detect the status of cervical lymph nodes is particularly needed clinically.

Method used

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  • LAMP primer group CK19-115 for quickly detecting lymphatic metastasis of head and neck squamous cell carcinoma
  • LAMP primer group CK19-115 for quickly detecting lymphatic metastasis of head and neck squamous cell carcinoma
  • LAMP primer group CK19-115 for quickly detecting lymphatic metastasis of head and neck squamous cell carcinoma

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0047] Example 1: collection and preservation of specimens

[0048] Fresh surgically resected specimens were collected, including head and neck tumor tissue, paracancerous tissue, normal pericancerous tissue, and cervical lymph node tissue. After the tissue specimens are taken, it is first necessary to determine the tumor location, extent, and infiltration that can be distinguished by the naked eye from the clinician, and describe and record them.

[0049] 1. Collection of tumor, paracancerous and pericancerous normal tissues:

[0050] Specimens were collected in accordance with the principle of "pericancer-adjacent cancer-tumor tissue". In order to ensure the mRNA activity of the specimen and determine the pathological characteristics of the tissue, the collected tissue specimens need to be processed as follows: 1) Take samples from the obvious parts of the tumor tissue (not necessarily the central part), avoiding the necrotic part; 2) Take a slightly larger cube or cuboid ...

Embodiment 2

[0064] Example 2: Screening of specific marker genes for HNSCC lymph node metastasis

[0065] 1. Identification of candidate genes

[0066] Possible mRNA markers for detection of HNSCC lymph node metastasis were identified by literature reports and database searches. Search sources include: PubMed, OMIM, UniGene, GeneCards, CGAP. The principle of pre-selection of candidate genes is that the expression of the gene in HNSCC is abnormal compared with normal lymph node tissue, that is, high expression or low expression, or the gene is up-regulated or down-regulated in HNSCC and has unique tissue specificity, or the gene has tumor-specific sex. Based on the above principles, a total of 39 candidate genes were pre-selected (see Table 1).

[0067] Table 1. Candidate genes for primary screening studies

[0068]

[0069] 2. qRT-PCR experiment to screen candidate genes twice

[0070] The specific experimental method is briefly described as follows:

[0071] 1) The first genetic...

Embodiment 3

[0121] Embodiment 3: Establishment of LAMP detection process

[0122] On the basis of determining the CK7, CK19 and CK7+CK19 gene combinations, we started to establish the LAMP experimental process. The establishment of the whole process includes three aspects: the preparation of the buffer solution of the experimental kit, the design of CK19 and CK7 primers, and the determination of the optimal reaction temperature. The core of which is the primer design. In the screening experiment of the best primers, the appropriate buffer solution and the best reaction temperature are obtained at the same time. 54 cases of tumor tissues from 36 HNSCC patients and 31 cases of benign lymph node tissues from 15 non-cancer patients were selected in the process establishment experiment, and the optimal primers and test reaction conditions were screened by establishing the RT-LAMP reaction system. The inclusion criteria of patients were the same as before.

[0123] 1) Primer design

[0124] ...

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Abstract

The invention screens out a specific marker gene and gene combination thereof for quickly detecting HNSCC lymphatic metastasis, namely CK7, CK19 and CK7+CK19, and establishes a lymph node quick detection technical system for an HNSCC patient, namely lymph node state detection by applying a RT-LAMP method, and the whole experiment process can be controlled in about 40 minutes. The detection method has the advantages of short time consumption, low experiment temperature, high result accuracy and simple and convenient interpretation mode. The HNSCC lymph node quick diagnosing system is established based on the RT-LAMP technology, and thus quick determination for the lymphatic metastasis state of patients during operation becomes possible.

Description

technical field [0001] The invention belongs to the field of life science and clinical medicine. More specifically, the present invention relates to a LAMP primer set for rapid detection of lymph node metastasis of head and neck squamous cell carcinoma, and a clinical, especially intraoperative rapid detection method for lymph node metastasis of head and neck squamous cell carcinoma. Background technique [0002] Head and neck squamous cell carcinoma (HNSCC) ranks sixth in the incidence of cancer in the world. There are nearly 650,000 new cases every year in the world. In recent years, the methods of diagnosis and treatment of malignant tumors are changing rapidly. However, the 5-year history of HNSCC Survival rates did not improve significantly. The choice of treatment options and prognosis of HNSCC mainly depend on TNM staging and histological grade, among which cervical lymph node metastasis has a greater impact on prognosis. Clinically, about 47% of HNSCC patients have...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6844C12Q1/6886C12Q2600/112C12Q2600/158C12Q2531/119C12Q2521/107
Inventor 邰隽倪鑫韩阳葛文彤张杰张亚梅王生才房居高陈晓红钟琦李平栋
Owner BEIJING CHILDRENS HOSPITAL AFFILIATED TO CAPITAL MEDICAL UNIV
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