Method for improving detection rate of cytokines secreted by peripheral blood leukocytes

A technology of cytokines and white blood cells, applied in the field of biomedicine, can solve the problems of low data and no statistical significance, and achieve the effect of a wide range of applications

Inactive Publication Date: 2018-11-16
WEIFANG MEDICAL UNIV
View PDF0 Cites 3 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, for some cytokines with relatively small secretion, such as IL-17, IL-4, IL-22, etc., because the final data ob

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Method for improving detection rate of cytokines secreted by peripheral blood leukocytes
  • Method for improving detection rate of cytokines secreted by peripheral blood leukocytes
  • Method for improving detection rate of cytokines secreted by peripheral blood leukocytes

Examples

Experimental program
Comparison scheme
Effect test

Embodiment

[0023] A method for improving the detection rate of cytokines secreted by peripheral blood leukocytes proposed by the present invention comprises the following steps:

[0024] S1. Take 500 μl whole blood and add 500 μl incomplete rpmi-1640 culture medium to dilute to obtain mixture A;

[0025] S2. Add phorbol lipid with a final concentration of 15 to 25 ng / ml and calciferin with a final concentration of 0.5 to 1.5 μg / ml to mixture A to obtain mixture B, and then place mixture B in a 24-well culture plate Cultivate for 6 hours, and add monensin at a final concentration of 2-3 μmol / L to block the secretion of cytokines in the last 4 hours of cultivation, and the cells to be tested are the cells after cultivation;

[0026] S3. Take 100 μl of cells to be tested after cultured in step S2, add CD3-PeCy5.5 and TCRγδ-PE, and stain in the dark at 4°C for 30 minutes, then dissolve red blood cells with whole blood samples, and then wash with 1ml staining buffer for 2 Once, add 500 μl of...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

No PUM Login to view more

Abstract

The invention discloses a method for improving the detection rate of cytokines secreted by peripheral blood leukocytes. The method comprises the following steps: S1, taking whole blood and adding a culture solution to dilute the whole blood; S2, adding phorbol ester and ionomycin calcium salt, carrying out culturing for 6 hours, and adding monensin in the later 4 hours of the culture so as to obtain cells to be detected; S3, adding a cell surface labeled antibody into the cells to be detected for staining of the cells to be detected, dissolving red blood cells with a whole blood sample, carrying out washing with a staining buffer, adding polyformaldehyde for lucifugal immobilization, adding a membrane permeating buffer for membrane permeation, adding bovine serum albumin and mouse serum for blocking, adding a cytokine antibody for staining, carrying out washing with membrane permeating buffer, and carrying out washing with a staining buffer to obtain stained cells; and S4, carrying outdetection with a flow cytometer. The method of the invention has a wide application range and high ratio of cytokine secreting cells, and experimental data and control data have statistical significance.

Description

technical field [0001] The invention relates to the technical field of biomedicine, in particular to a method for improving the detection rate of cytokines secreted by peripheral blood leukocytes. Background technique [0002] Flow cytometry is a common method used by most researchers to detect secreted cytokines in peripheral blood samples. In experiments, it is customary to separate mononuclear cells (PBMC) in peripheral blood samples or directly sort T cell subsets for detection. However, for some cytokines with relatively small secretion, such as IL-17, IL-4, IL-22, etc., because the final data obtained are relatively low, many experimental data are not statistically significant when compared with the control group. Because the secretion of one cytokine requires the assistance of other cytokines, and there are many cytokines in serum, as reported by Manel N et al., in the absence of serum, initial CD4+ T cells secrete IL-17 dependent on TGF- Presence of β, IL-1β, IL-6,...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
IPC IPC(8): G01N33/68G01N15/14
CPCG01N15/14G01N33/6863
Inventor 彭美玉
Owner WEIFANG MEDICAL UNIV
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap