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Method and kit for separation and purification of fetal nucleated red blood cells

A technology for separation and purification of nucleated erythrocytes, applied in the field of prenatal gene or chromatin inspection, isolation and purification methods and kits for fetal nucleated erythrocytes, which can solve the problem of fetal cell enrichment, small number of separations, and no substantial breakthroughs , limited range of diseases, etc., to achieve the effect of tolerating separation and washing operations, avoiding a large amount of loss, and improving purity

Active Publication Date: 2020-04-10
SHANDONG YADA PHARMA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] To sum up, the current domestic and foreign researches in this area all have the problems of enrichment of fetal cells, small number of separations, insufficient purity, complex operation process, and the scope of diseases applied to prenatal diagnosis is very limited, making this work insubstantial. sexual breakthrough

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0065] Embodiment 1: Separation and purification of fetal NRBC

[0066] Take 3ml of peripheral venous blood from pregnant women whose pregnancy is 16-18 weeks into heparin anticoagulant blood collection tubes, centrifuge at 500g for 5 minutes to remove plasma, and add 8ml of PBS (NaCl 137mmol / L, KCl 2.7mmol / L, NaCl 2.7mmol / L, NaCl 2 HPO 4 4.3mmol / L, KH 2 PO 4 1.4mmol / L, pH7.2) prepared 1% (w / v) NH 4 Cl solution, mix well, place in ice bath for 10 minutes, lyse red blood cells, centrifuge at 500g for 5 minutes to remove supernatant, shake the pellet to resuspend the cells. Then add 1ml of 3% glutaraldehyde solution prepared in PBS, and fix the cells for 10 minutes. Fully wash the cells twice with 10 ml of normal saline, resuspend the cells with 1 ml of normal saline, add 10E+8CD45 antibody-labeled magnetic beads (Miltenyl biotech, serial number 130-045-801), mix well for 15 minutes, and place on a magnetic separation rack Separate the cells, transfer the liquid part to a...

Embodiment 2

[0068] Take 3ml of peripheral venous blood from a pregnant woman whose pregnancy is 16-18 weeks into a heparin anticoagulant blood collection tube, centrifuge at 500g for 5 minutes to remove plasma, add 8ml of 1% NH prepared in PBS to the blood cell pellet 4 Cl solution, mix well, place in ice bath for 10 minutes, lyse red blood cells, centrifuge at 500g for 5 minutes to remove supernatant, shake the pellet to resuspend the cells. Then add 1ml of 3% glutaraldehyde solution prepared in PBS to fix the cells for 10 minutes. Fully wash the cells twice with 10 ml of normal saline, resuspend the cells with 1 ml of normal saline, add 10E+8CD45 antibody-labeled magnetic beads (Miltenyl biotech, serial number 130-045-801), mix well for 15 minutes, and place on a magnetic separation rack Separate the cells, transfer the liquid part to a test tube containing 10E+6CD71 antibody-labeled magnetic beads (Miltenyl biotech, serial number 130-046-201), mix well for 15 minutes, separate the cell...

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Abstract

The invention relates to a method for separating and purifying fetal nucleated erythrocytes, which is characterized by comprising the following steps: lysing the anucleated erythrocytes in the peripheral venous blood samples of pregnant women; adding a cell immobilization reagent to the lysed blood samples; and Cell separation is performed on a cell-fixed blood sample to obtain fetal nucleated red blood cells. The invention also relates to a kit for separating and purifying fetal nucleated red blood cells from peripheral venous blood samples of pregnant women. The invention greatly improves the yield and purity of isolated target cells, and lays a good foundation for subsequent various analyses.

Description

technical field [0001] The invention belongs to the field of cell biology detection, and in particular relates to a separation and purification method and kit for fetal nucleated red blood cells. The invention can be applied to prenatal genetic or chromatin screening. Background technique [0002] With the development of modern medicine, the advancement of medical technology, and the development of medical and health care, infectious diseases have been better controlled, and people's average life expectancy has been continuously extended. There is no effective treatment, and it is becoming more and more important in the diseases that seriously threaten people's health, which has brought great mental burden and economic loss to the family and society. Therefore, prenatal diagnosis is carried out to avoid the birth of children as a control genetic disease. It is an important way to prevent the occurrence of diseases, reduce the genetic load of the population, and improve the ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/073C12N5/078
CPCC12N5/0641C12N2509/00C12N5/00G01N33/68
Inventor 夏庆杰杨林杨雪飞
Owner SHANDONG YADA PHARMA
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