Separating and purifying method for ox adrenal medella chromaffin cell

A technology for separation and purification of chromaffin cells, applied in the field of separation and purification of bovine adrenal medulla chromaffin cells, which can solve the problems of long duration, unclear cell level, and increased process costs, so as to simplify the operation process and improve cell harvest Effect of quantity, source of healthy cells

Inactive Publication Date: 2005-08-03
DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
View PDF3 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] ①Using collagenase digestion as the main cell separation method, the large-scale use of high-concentration collagenase and long digestion time not only lead to a decrease in cell yield, but also greatly increase the process cost;
[0005] ②Because of the heterogeneity of chromaffin cells and the distribution of cell density within a certain range, the continuous density gradient centrifugation method to purify chromaffin cells will not only cause loss when the cells are collected, but also the cell level is not clear enough after ce

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
  • Separating and purifying method for ox adrenal medella chromaffin cell
  • Separating and purifying method for ox adrenal medella chromaffin cell
  • Separating and purifying method for ox adrenal medella chromaffin cell

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] Isolation of bovine adrenal medulla chromaffin cells

[0022] 1. Select the adrenal glands of young bulls whose warm ischemia time is less than 30min from the slaughterhouse, store them in D-Hank's (containing 10000units / ml of penicillin and streptomycin) solution at 4°C, place them on ice and bring them back to the laboratory, use The method of the invention can simultaneously complete the separation and purification of 8-12 bovine adrenal chromaffin cells.

[0023] 2. After infusing calcium- and magnesium-free D-Hank's solution retrogradely, soak the adrenal gland in D-Hank's solution to fully ooze out the blood cells remaining in the blood vessels.

[0024] 3. Intravenous infusion of 0.1% type I collagenase 5ml against the adrenal gland, digested in a constant temperature shaking water bath at 37°C for 20 minutes, to fully dissociate the collagen components between the cortex, medulla and medulla cells.

[0025] 4. Peel off the cortex and cut the medulla into 1mm as...

Embodiment 2

[0032] Example 2: The isolated and purified cells were subjected to glyoxylic acid-induced characteristic fluorescent staining of catecholamines, and the yellow-green fluorescence of chromaffin cells could be observed, confirming that the isolated and purified cells are cells with the characteristics of secreting catecholamines—namely, chromaffin cells cells (see figure 2 ).

Embodiment 3

[0033] Example 3: 7 ml of type I collagenase with a mass percentage concentration of 0.05% was infused into the reverse adrenal gland vein, and digested in a 37°C constant temperature shaking water bath for 50 minutes to fully dissociate the collagen components between the cortex, the medulla and the medulla cells, All the other conditions are with embodiment 1.

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

PropertyMeasurementUnit
Densityaaaaaaaaaa
Diameteraaaaaaaaaa
Login to view more

Abstract

The ox adrenal medella chromaffin cell separating and purifying method includes perfusing collagenase solution backwards to adrenal vein; fully hydrolyzing adrenal medella cell and collagen between cortical cell and medella cell under certain temperature; mechanically shearing and gradiently sieving stripped medella to obtain suspension with discrete adrenal chromaffin cell; and purifying the cell by means of the difference in cell density in the suspension. In the Percoll non-continuous gradient method to purifying ox adrenal medella chromaffin cell, every adrenal can be separated to obtain (0.7-3.1)E8 chromaffin cells of purity over 95 % and with excellent shape and activity over 99 % averagely.

Description

technical field [0001] The invention relates to a method for separating and purifying bovine adrenal medulla chromaffin cells. Background technique [0002] Adrenal medulla chromaffin cells are neuronal cells that synthesize, store and secrete catecholamines. When implanted in the central nervous system, it can remain active for a long time and secrete catecholamines on demand. Although a large number of in vivo tests have also shown its significant effect in alleviating pain and treating Parkinson's disease, the technology is still in the treatment stage of individual clinical cases. There are still some problems in its wide application in clinical practice, such as limited cell sources, adrenal chromaffin cells cannot divide and proliferate in vitro, and there is no cell line with good functional expression, and the cells can only be obtained by separating the adrenal medulla; The separation and purification process of chromaffin cells reported in the past is complicated...

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): C12N5/071
Inventor 马小军张晓慧王为
Owner DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
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