Kit for colorectal cancer auxiliary diagnosis and/or prognosis

A technology for auxiliary diagnosis and prognosis judgment, applied in biological testing, biochemical equipment and methods, microbial measurement/testing, etc., can solve problems such as blocking tumor formation

Inactive Publication Date: 2015-09-02
黄文林
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Some tumor studies have reported the function of PCDH family molecules in cancer. For example, in Wilm's tumor, all PCDH genes were found to be hypermethylated; PCDH10 was silenced due to hypermethylation in many tumors, which is considered to be A potential tumor suppressor gene; PCDH24 can cause cell contact inhibition in the colorectal cancer HCT116 cell line, thereby completely blocking tumor formation in vitro, and this effect may be achieved by negatively regulating the Wnt pathway, but currently There is no report about PCDHB3

Method used

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  • Kit for colorectal cancer auxiliary diagnosis and/or prognosis
  • Kit for colorectal cancer auxiliary diagnosis and/or prognosis
  • Kit for colorectal cancer auxiliary diagnosis and/or prognosis

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0055] Embodiment 1, clinical sample collection

[0056] For the determination of mRNA content: a total of 98 colorectal cancer samples from February 2010 to August 2012 were selected from Sun Yat-Sen University Cancer Center, and fresh tissues were taken after surgery and stored in liquid nitrogen; for protein content determination samples: select A total of 210 colorectal cancer specimens were collected from Sun Yat-sen University Cancer Center from September 2000 to January 2006. The paraffin specimens were stored at room temperature, and the cut white slices were stored in a 4°C refrigerator.

[0057] For each selected case, the inclusion criteria were newly diagnosed, pathologically diagnosed as single colorectal adenocarcinoma, no preoperative radiotherapy and chemotherapy, and radical resection was required for TNM stage I, II, and III cases. Each patient contained tumors and normal controls (normal mucosa more than 5 cm from the resection margin).

[0058] For all cas...

Embodiment 2

[0059] Example 2, quantitative PCR detection of PCDHB3 mRNA levels in colorectal cancer tissues and cell lines

[0060] 1) Total RNA was extracted from 98 colorectal cancer tissues and their normal controls, 8 colorectal cancer cell lines and two normal control cell lines using the TRIZOL method, and 2 μg of total RNA was reverse-transcribed into cDNA.

[0061] 2) Query the Gene database of NCBI to obtain the cDNA sequences of PCDHB3 and GAPDH (internal reference), and design PCR primers according to the sequences as follows:

[0062]

[0063] 3) Establish the following reaction system, and set up three reaction wells for each sample:

[0064]

[0065] Mix the above components, add to each well of the 8-strip tube, close the lid tightly, remove air bubbles, and centrifuge to make the liquid gather to the bottom of the tube. The thermal cycle parameters were as follows: 95°C for 10 min, 95°C for 30 s, 60°C for 30 s, 72°C for 40 s, 40 cycles, 95°C for 1 min, 60°C for 30 s...

Embodiment 3

[0070] Example 3, Determination of PCDHB3 protein content in colorectal cancer tissue specimens

[0071] 1) During the experiment, 210 cases of colorectal cancer cases were taken out paraffin section white slices, dewaxed with xylene, hydrated with gradient alcohol, and 0.3% H 2 o 2 Detissue peroxidase, citrate solution microwave repair, Abgent anti-PCDHB3 (AP12159a) antibody (1:100) and corresponding species secondary antibody incubation, DAB and hematoxylin color development, hydrochloric acid alcohol differentiation, gradient alcohol dehydration After the xylene transparency step, the slides were sealed with neutral gum.

[0072] 2) The staining evaluation of tissue samples was taken by a 200× inverted microscope, and 5 visual fields / cases were taken for each of the paracancerous and cancerous tissues, and then the pictures were analyzed with inForm1.4.0 software. The staining intensity of PCDHB3 in cells was divided into four grades: negative (0), weak positive (1), mode...

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Abstract

The invention discloses a kit for colorectal cancer auxiliary diagnosis and / or prognosis. The kit for colorectal cancer auxiliary diagnosis and / or prognosis is developed based on a gene PCDHB3 related to colorectal cancer. The kit comprises at least one of the following reagents of a primer pair for detecting the PCDHB3 gene expression level, a primer pair for detecting the PCDHB3 gene methylation level and the reagent for detecting the PCDHB3 protein level. Differential expression gene screening is conducted on the colorectal cancer, and PCDHB3 located in human body chromosomes 5q31 is the gene closely related to the colorectal cancer. The mRNA and protein expression level of the PCDHB3 is remarkably decreased in the tumor tissue of a colorectal cancer patient, and the decrease is closely related to gene promoter region CpG basic group height methylation. Meanwhile, prognostic analysis of the TNM III-IV stage colorectal cancer patient shows that the PCDHB3 indicates good prognosis in tumor cell high-level expression or tumor cell karyotheca expression.

Description

technical field [0001] The invention belongs to the field of molecular diagnosis, and specifically relates to a kit for auxiliary diagnosis and / or prognosis judgment of colorectal cancer developed based on the gene PCDHB3 related to colorectal cancer. Background technique [0002] Currently, colorectal cancer is one of the most common tumors worldwide, with more than one million new cases and more than 600,000 deaths each year. In China, the mortality rate of colorectal cancer is 7.25 / 100,000, second only to lung cancer, liver cancer, gastric cancer and esophageal cancer. In recent years, with the improvement of people's living standards in our country, changes in dietary structure, lack of exercise, smoking and other factors, the incidence of colorectal cancer is on the rise, especially in large and medium-sized cities with developed coastal economies, the incidence of colorectal cancer has reached It even surpasses the average level of western developed countries, becomes...

Claims

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Application Information

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IPC IPC(8): C12Q1/68G01N33/68G01N33/574
CPCC12Q1/6886C12Q2600/118C12Q2600/154C12Q2600/158G01N33/574G01N33/57446G01N33/68G01N2333/705
Inventor 黄文林叶文
Owner 黄文林
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