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Application of CD11b positive exosome in tumor prognosis evaluation

A prognostic assessment and exosome technology, applied in the field of biomedicine, can solve the problems of reduced specificity and sensitivity, the relationship between S100A9 protein and colon cancer recurrence and metastasis has not been found, and achieves high repeatability, good specificity and easy operation. Effect

Pending Publication Date: 2021-10-19
JIANGSU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Serum S100A9 has high sensitivity and specificity in the diagnosis of colorectal cancer, but its specificity and sensitivity are greatly reduced in the diagnosis of colorectal cancer metastasis, only about 50%
There is no report on the relationship between S100A9 protein in exosomes and the recurrence and metastasis of colon cancer

Method used

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  • Application of CD11b positive exosome in tumor prognosis evaluation
  • Application of CD11b positive exosome in tumor prognosis evaluation
  • Application of CD11b positive exosome in tumor prognosis evaluation

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0018] Example 1: CD11b in plasma of patients with colorectal cancer + Determination of exosomes content

[0019] (1) Preparation of plasma exosomes:

[0020] Peripheral blood samples collected from healthy volunteers (40 cases) and colorectal cancer patients (72) were placed at room temperature for 30 minutes, then centrifuged at 1200g for 10 minutes at 4°C to separate plasma. Plasma was centrifuged at 1800 g for 10 min at 4°C, wrapped in cotton and stored at -80°C for later use.

[0021] ②Take out the preserved plasma sample and put it on ice to thaw, and centrifuge at 1500 g for 10 min at 4°C. After collecting the supernatant, centrifuge at 10,000 g for 20 min at 4°C to collect the supernatant.

[0022] Add 2.5 μL thrombin to 250 μL supernatant and incubate at room temperature for 5 min. Centrifuge at 10,000 g for 5 min at 4°C to collect the supernatant; add the ExoQuick-TCTM Exosomes reagent (5:1) in the SBI ExoQuick-TCTM Kit (Product No.: EXOQ5A-1) to the supernat...

Embodiment 2

[0029] Example 2: CD11b in plasma of patients with colorectal cancer + exosomes and CD11b - Determination of S100A9 content in exosomes

[0030] Sample preparation: CD11b was prepared by the same method as in Example 1 - Colloidal particles loaded with exosomes, respectively, CD11b + Colloidal particles loaded with exosomes and CD11b - The colloidal particles loaded by exosomes were mixed with an equal volume of lysate (RIPA: PMSF = 250:1), placed on ice for 1 h, shaken once every 10 min; centrifuged at 12000 g for 15 min at 4 °C, and the supernatant was collected.

[0031] The protein concentration in the exosomes lysate was determined by BCA microprotein quantification using the BCA Protein Assay Kit (Cat. No.: CW0014S).

[0032] Take out the required strips from the aluminum foil bag after equilibrating at room temperature for 20 min, and seal the remaining strips with a ziplock bag and return them to 4°C.

[0033] Set standard wells and sample wells, add 50 ...

Embodiment 3

[0041] Example 3: Plasma CD11b of colorectal cancer patients and healthy controls before and after surgery + Determination of exosomal S100A9 levels

[0042] In this example, the peripheral blood samples collected from 72 patients with colorectal cancer were collected; among them, 46 patients with colorectal cancer without surgery (CoCa) and 22 patients with colorectal cancer without recurrence after surgery (SuFr). Peripheral blood samples from 4 patients with recurrent colorectal cancer (SuRe) and 40 healthy controls (Ctrl) were tested for plasma CD11b + Detection of exosomal S100A9 content.

[0043] Sample preparation: lyse CD11b in colorectal cancer patients and healthy controls, respectively + Exosomes, using the BCAProtein Assay Kit to quantify the protein concentration by BCA Microprotein.

[0044] According to the instructions of the RD catalog S100A9 detection kit (catalog number: DS8900), a standard curve was prepared, and the content of S100A9 in each sample...

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Abstract

The invention belongs to the technical field of biological medicines, and particularly relates to an application of a CD11b positive exosome in tumor prognosis evaluation. According to the invention, peripheral blood samples of healthy people, colorectal cancer patients and patients before and after an operation are collected, human plasma CD11b + exosomes and CD11b-exosomes are sorted by adopting flow cytometry, and the content of S100A9 in the CD11b + exosomes is detected by utilizing ELISA. The S100A9 protein in the plasma CD11b + exosomes is determined to have a significant high expression quantity, and has sensitivity and specificity for judging recurrence and metastasis of a colorectal cancer; and meanwhile, the application has the characteristics of simple operation, sensitive detection, good specificity, high repeatability and the like. The application confirms that the expression level of the S100A9 protein in CD11b + exosomes is related to the prognosis of the postoperative colorectal cancer patient, can be used for judging the prognosis of the colorectal cancer patient, and has important guiding significance for reducing the death rate of the colorectal cancer and improving the prognosis.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and in particular relates to the application of CD11b-positive exosomes in tumor prognosis assessment. Background technique [0002] Colorectal cancer is one of the most common digestive tract malignancies with a high mortality rate. Although there are many markers for the diagnosis of colorectal cancer, there is a lack of detection methods and biomarkers involved in prompting postoperative recurrence and metastasis. Serum S100A9 has high sensitivity and specificity in the diagnosis of colorectal cancer, but its specificity and sensitivity are greatly reduced in the diagnosis of colorectal cancer metastasis, only about 50%. [0003] Exosomes are membranous vesicles with a diameter of 30-150 nm that contain proteins and nucleic acids from the cell of origin. The contents of exosomes can be transferred from source cells to target cells, playing an important role in intercellular information t...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N15/14
CPCG01N15/14G01N2015/1006G01N15/149
Inventor 查璇魏来王胜军
Owner JIANGSU UNIV
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