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Separation and purification method for foetus erythroblast in maternal blood and separator

A kind of technology for pregnant women's peripheral blood and separation methods, which is applied in the direction of cell dissociation methods, blood/immune system cells, biochemical equipment and methods, etc., which can solve the problems of cumbersome operation steps, low sensitivity, and long time consumption, and achieve simple operation steps , High sensitivity, reduce the effect of hemolysis

Inactive Publication Date: 2020-06-05
SHANDONG PROVINCIAL HOSPITAL +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The existing technical system has problems such as low capture efficiency, high cost, cumbersome operation steps, long time-consuming, low sensitivity and poor specificity, which limit its clinical application

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] A method for isolating nucleated erythrocytes from fetal blood in pregnant women, the specific steps are as follows:

[0035] (1) CD71-positive cell line (K562) culture and antibody binding ability verification;

[0036] ① Cell culture: CD71 positive cell line (K562) culture;

[0037] ② Immunofluorescence technology to verify the binding ability of CD71 antibody: select CD71 polyclonal antibody (ab8403), CD71 monoclonal antibody (ab214039), CD45 mouse monoclonal antibody (ab8216), CD147 polyclonal antibody (ab64616), goat polyclonal antibody secondary antibody To mouse (ab150116), goat polyclonal antibody secondary antibody to rabbit (ab150077) for immunofluorescence experiments to verify the binding ability of the antibody;

[0038] (2) Establish a capture system

[0039] ③ Type and diameter of microspheres: make 1μm ferric oxide microspheres;

[0040] ④ Use K562 cells to obtain the best capture conditions: 1 μm ferric oxide microspheres are coupled with CD71 polycl...

Embodiment 2

[0049] A method for isolating nucleated erythrocytes from fetal blood in pregnant women, the specific steps are as follows:

[0050] (1) CD71-positive cell line (K562) culture and antibody binding ability verification;

[0051] ① Cell culture: CD71 positive cell line (K562) culture;

[0052] ② Immunofluorescence technology to verify the binding ability of CD71 antibody: select CD71 polyclonal antibody (ab8403), CD71 monoclonal antibody (ab214039), CD45 mouse monoclonal antibody (ab8216), CD147 polyclonal antibody (ab64616), goat polyclonal antibody secondary antibody To mouse (ab150116), goat polyclonal antibody secondary antibody to rabbit (ab150077) for immunofluorescence experiments to verify the binding ability of the antibody;

[0053] (2) Establish a capture system

[0054] ③Types and diameters of microspheres: make 1μm ferric oxide microspheres;

[0055] ④ Use K562 cells to obtain the best capture conditions: 1 μm ferric oxide microspheres are coupled with CD71 polyc...

Embodiment 3

[0064] A method for isolating nucleated erythrocytes from fetal blood in pregnant women, the specific steps are as follows:

[0065] (1) CD71-positive cell line (K562) culture and antibody binding ability verification;

[0066] ① Cell culture: CD71 positive cell line (K562) culture;

[0067] ② Immunofluorescence technology to verify the binding ability of CD71 antibody: select CD71 polyclonal antibody (ab8403), CD71 monoclonal antibody (ab214039), CD45 mouse monoclonal antibody (ab8216), CD147 polyclonal antibody (ab64616), goat polyclonal antibody secondary antibody To mouse (ab150116), goat polyclonal antibody secondary antibody to rabbit (ab150077) for immunofluorescence experiments to verify the binding ability of the antibody;

[0068] (2) Establish a capture system

[0069] ③ Type and diameter of microspheres: make 1μm ferric oxide microspheres;

[0070] ④ Use K562 cells to obtain the best capture conditions: 1 μm ferric oxide microspheres are coupled with CD71 polycl...

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PUM

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Abstract

The invention belongs to the field of cell biology detection, and particularly relates to a separation and purification method for foetus erythroblast in maternal blood and a separator. The method comprises the steps of (1) culturing a CD71 positive cell line (K562) and detecting antigen binding capacity and specificity; (2) establishing a catching system by a CTCBIOPSY-detection system and K562 cells and optimizing capturing conditions by cord blood; and (3) separating the collected foetus erythroblast: enlarging the diameter of target cells by microspheres coupled with a CD71 polyclonal antibody, and through the CTCBIOPSY- detection system, separating the erythroblast in the maternal blood by a filtering principle. The method is high in capturing efficiency, low in cost, simsple in operation steps, short in consumption time and high in sensitivity.

Description

technical field [0001] The invention belongs to the field of cell biology detection, and in particular relates to a method for separating and purifying fetal nucleated red blood cells and a separator. Background technique [0002] Birth defects refer to a general term for abnormalities in body structure, function or metabolism that occur before a baby is born, usually including congenital structural deformities, chromosomal abnormalities, genetic metabolic diseases, and functional abnormalities such as blindness, deafness, and mental retardation. In September 2012, the "Report on the Prevention and Control of Birth Defects in China" issued by the former Ministry of Health pointed out that the current incidence of birth defects in China is about 5.6%, and the number of new birth defects is about 900,000 cases every year. There are about 250,000 cases of birth defects. Birth defects are the main cause of early miscarriage, premature birth, stillbirth, perinatal death, and inf...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/078
CPCC12N5/0641C12N2509/00
Inventor 韩波李胜冯鑫鑫张丽李浩黄劭毅姜殿东王静吕建利
Owner SHANDONG PROVINCIAL HOSPITAL
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