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A method for isolating placental trophoblast cells from exfoliated cells of the cervix of pregnant women

A technology of trophoblast cells and exfoliated cells, which is applied in the field of separation and screening of trophoblast cells, can solve problems such as limited antibody combinations, and achieve the effects of early sampling time, good specificity, and low risk of infection and abortion

Active Publication Date: 2021-08-24
GUANGZHOU HYBRIBIO MEDICINE TECH LTD +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

On the other hand, there is still room for improvement in this technology. The disclosed antibody combinations are limited, and the accuracy and specificity need to be improved.

Method used

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  • A method for isolating placental trophoblast cells from exfoliated cells of the cervix of pregnant women
  • A method for isolating placental trophoblast cells from exfoliated cells of the cervix of pregnant women
  • A method for isolating placental trophoblast cells from exfoliated cells of the cervix of pregnant women

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0047] Example 1 Microfluidic Sorting Chip Design

[0048] Schematic diagram of the microfluidic sorting chip used to separate and screen trophoblast cells from the exfoliated cells of the cervix of pregnant women. figure 1 shown.

[0049] The structural design of the microfluidic sorting chip is described as follows: the chip is prepared using but not limited to acrylic as the basic material, and is formed on one side of the substrate by injection molding technology. figure 1 For the shape of the pipe, the width of the pipe does not exceed 1000 μm, and the depth does not exceed 500 μm, and the other side of the substrate is used for lamination to form a complete chip.

[0050] Specifically, the microfluidic sorting chip includes a substrate and a cover attached thereto;

[0051] Such as figure 1 As shown, one side of the substrate is provided with a main channel, a side channel A and a side channel B, and the two side channels are respectively close to the left and right end...

Embodiment 2

[0057] Example 2 Separation and screening of trophoblast cells from cervical exfoliated cells of pregnant women based on microfluidic sorting chip

[0058] One, the sorting method of trophoblast cell comprises the following steps:

[0059] 1. Prepare the cervical exfoliated cell fluid sample into a sample cell suspension; the specific methods include as shown in (1)-(5) below;

[0060] 2. Add specific antibodies to the sample cell suspension for incubation; specific methods include as shown in (6)-(14) below;

[0061] 3. Using the microfluidic sorting chip of Example 1, the cell suspension incubated in step 2 was subjected to fluorescence-labeled microfluidic cell sorting; specific methods include as shown in (15)-(18) below.

[0062] Wherein, the combination of specific antibodies is shown in Table 1:

[0063] Table 1: Combinations of antigens and antibodies expressed on trophoblast cells

[0064]

[0065] Two, specifically, the sorting method of trophoblast cells compr...

Embodiment 3

[0084] Example 3 Method for Separating and Screening Trophoblast Cells from Pregnant Women's Cervical Exfoliated Cells Based on Flow Cytometry

[0085] The sorting method of trophoblast cells comprises the following steps:

[0086] 1. The cervical exfoliated cell liquid sample is prepared into a sample cell suspension; the specific method is shown in (1)-(5) in Example 2;

[0087] 2. Add specific antibodies to the sample cell suspension and incubate; the specific method is the same as shown in (6)-(14) in Example 2; wherein, the combination of specific antibodies is the same as shown in the above table 1;

[0088] 3. Use a flow cytometry sorter (BDFACSAria II, USA) to perform fluorescent marker sorting on the cell resuspension incubated in step 2, including the following steps:

[0089] 1) Turn on the flow cytometer, and follow the operating instructions for daily start-up operations;

[0090] 2) Adjust the liquid flow of the instrument so that the breakpoint of the liquid f...

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Abstract

The invention discloses a method for separating placental trophoblast cells from exfoliated cells of the cervix of pregnant women. The method is based on the specific antigens and combinations expressed on the surface or intracellular surface of trophoblast cells determined by research, and uses the designed microfluidic sorting chip or flow cytometer to sort the cell suspension of placental trophoblast samples , to obtain isolated and purified placental trophoblast cells. Compared with the traditional method, the present invention has the advantages of non-invasive acquisition of specimens and good specificity, and the method takes earlier time, which leads to a lower risk of infection and abortion, and can realize synchronous labeling and characteristic fluorescence signals of multiple antigens at the same time The accuracy of identification and sorting has been greatly improved, and the detection results have higher reliability and wider coverage.

Description

technical field [0001] The invention belongs to the technical field of cell separation. More specifically, it relates to a method for separating and screening trophoblast cells from exfoliated cells of the cervix of pregnant women. Background technique [0002] The pre-pregnancy, prenatal and neonatal three-level prevention and control system is one of the important means to reduce birth defects and improve the quality of the population in my country. Among them, prenatal screening and diagnosis are the most complicated and difficult links. [0003] At present, amniocentesis or umbilical vein puncture used clinically for prenatal diagnosis, and non-invasive prenatal screening technologies such as fetal cell-free nucleic acid sequencing have various deficiencies; The risk of abortion and miscarriage is high, the analysis time is long, the detection items and scope are limited, and the diagnosis time is limited to the second and third trimesters, the acceptance of pregnant wo...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/073G01N15/14
CPCC12N5/0605G01N15/14C12N2509/10G01N2015/1006G01N15/1459C12N5/0081G01N33/56966C07K16/18C07K16/28C07K16/26C07K16/2803C07K16/40G01N15/149G01N33/533G01N33/5005C12Q1/68
Inventor 郑焱马佳艳丰晓威刘梦羽陈佩璇管秩生谢龙旭葛毅媛
Owner GUANGZHOU HYBRIBIO MEDICINE TECH LTD
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