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Application of a set of tumor-associated antigens in early screening kit for esophageal cancer and kit thereof

A tumor-associated antigen, esophageal cancer technology, applied in the field of tumor molecular biology, can solve the problems of limitation, low efficiency, high cost, etc., and achieve the effects of good clinical application prospects, convenient use and small side effects

Inactive Publication Date: 2019-09-03
THE FIRST AFFILIATED HOSPITAL OF ZHENGZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, traditional chromoendoscopy uses iodine, but the use of iodine may cause some adverse reactions to the human body; endoscopic narrow-band imaging technology uses electronic dyeing, which has no side effects of chemical dyes, but is expensive and traumatic in endoscopic examination , the efficiency is low, thus limiting the promotion of endoscopic narrow-band imaging examination

Method used

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  • Application of a set of tumor-associated antigens in early screening kit for esophageal cancer and kit thereof
  • Application of a set of tumor-associated antigens in early screening kit for esophageal cancer and kit thereof
  • Application of a set of tumor-associated antigens in early screening kit for esophageal cancer and kit thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0025] Example 1: Discovery and identification of biomarkers in esophageal cancer tumors

[0026] (1) Protein extraction from patient biopsies

[0027] 500 mg of patient paired biopsies (tumor biopsies versus adjacent normal tissue) were collected and washed with PBS, snap-frozen by immersion in liquid nitrogen, then removed, homogenized using a pestle and mortar, and injected into Add 5 µL lysis solution to the homogenized sample, which contains 8mol / L urea, 4% (v / v) CHAPS, 2% (v / v) IPG buffer and 0.2mg / mL PMSF, then vortex to mix At least 5min until the tissue is completely dispersed; then centrifuge at 14000rpm / min for 10min at 4°C, centrifuge the lysate to clarify, and take the supernatant for later use; use the cleaning kit, namely 2D CleanUp kit, to further clean the supernatant to remove salt and impurities; resuspend the pellet with 1 µL of rehydration solution (lysed solution without DTT&IPG buffer); then use the Bio-Rad protein assay to measure the concentration of ...

Embodiment 2

[0036] Example 2: In vitro expression and protein purification of biomarker panels

[0037] (1) In vitro expression of biomarker panels

[0038] Based on the amino acid sequence of the targeted biomarkers, commercially synthesized cDNA clones are used to express the biomarker panel. The in vitro expression process of the biomarker panel is as follows: figure 2 As shown, the specific process is as follows:

[0039] HIS-tagged plasmids containing cDNA inserts encoding biomarker panels were transformed into DH5 competent cells, single colonies were picked and grown in bacterial culture, the number of plasmids was expanded, and extracted from bacteria by miniprep Extract the plasmid, further transform the plasmid into BL21DE3 or BL21DE3pLysS competent cells, select the transformed bacteria and grow them in 2X 100mL LB medium, when the bacterial culture reaches an OD value of 0.06, add 200µM to 100mL bacterial culture For IPTG, use 100 mL of bacterial culture without IPTG as a neg...

Embodiment 3

[0044] Example 3: Preparation of biomarker-BioPlex bead conjugates

[0045] The biomarker histone prepared by referring to the method described in Example 2 or the purchased biomarker histone with a purity of more than 90% were used to couple the biomarker histone to the BioPlex beads. The specific process was to combine the uncoupled The linked BioPlex beads were vortexed for 30s, then sonicated for 15s, and 1,250,000 BioPlex beads were collected in a reaction tube by centrifuging 100µL BioPlex beads at 10000rpm / min for 4min, and washed with 100µL BioPlex bead washing buffer by centrifugation Finally, resuspend the BioPlex beads in 80 µL of BioPlex bead activation buffer, which is MES buffer containing EDC / NHS activation system, and then add 10 µL of 50 mg / mL freshly prepared N-hydroxysulfosuccinate to the BioPlex beads Imine (S-NHS) aqueous solution and 10 µL of 50 mg / mL freshly prepared 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) aqueous solution were incubated at ...

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Abstract

The invention belongs to the field of tumor molecular biology, and particularly relates to application of a set of tumor-associated antigens in an early screening kit for esophageal cancer and a kit thereof. The kit provided by the invention comprises a solid phase carrier and esophageal cancer tumor-associated antigens coated on the solid phase carrier, wherein the esophageal cancer tumor-associated antigens consists of a CP96 gene expression, a CRP78 gene expression, an MMP-10 gene expression, a cav-1 gene expression, a CDC25B gene expression, a PGK1 gene expression and an HSP105 gene expression; and the kit further comprises a phycoerythrin-conjugated secondary antibody. The detection specificity and sensitivity of the kit provided by the invention for esophageal cancer are as high as 96.1% and 95.1%, respectively. Compared with the tissue biopsy, the kit provided by the invention only needs to collect the blood of the person to be examined to screen the risk of esophagus cancer with small trauma and side effects, and is easy to use and provided with good clinical application prospects.

Description

technical field [0001] The invention belongs to the field of tumor molecular biology, and specifically relates to the application of a group of tumor-associated antigens in an early screening kit for esophageal cancer and the kit. Background technique [0002] Esophageal cancer is one of the six most common malignant tumors in the world, and its morbidity and mortality rate occupy the sixth place; China is the country with the highest incidence rate and death rate of esophageal cancer, the national incidence rate ranks sixth, and the mortality rate ranks 1st fourth place. Of the 500,000 new cases of esophageal cancer each year worldwide, more than half occur in China. Significantly different from the Western population, the prominent epidemiological features of Chinese esophageal cancer are significant regional distribution differences and obvious familial clustering. The Taihang Mountains at the junction of Henan, Hebei, and Shanxi provinces, Linzhou, Henan is the area wi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/574
CPCG01N33/57407
Inventor 王立东韩月霞宋昕赵学科王盼盼周英发袁翎
Owner THE FIRST AFFILIATED HOSPITAL OF ZHENGZHOU UNIV
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