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A method for in situ culture of amniocytes and karyotype processing and analysis on glass slides

A technology of amniotic fluid cells and in situ culture, which is applied to animal cells, vertebrate cells, artificial cell constructs, etc., can solve the problems of high analysis cost and difficult popularization, and achieve high-quality cleavage phase, stable production quality, and high success rate high effect

Active Publication Date: 2017-10-20
WENZHOU CENT HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Tabor et al. used slide culture dishes from NUNC Company for in situ culture of amniocytes, but the analysis cost is too high and it is not easy to popularize in China

Method used

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  • A method for in situ culture of amniocytes and karyotype processing and analysis on glass slides
  • A method for in situ culture of amniocytes and karyotype processing and analysis on glass slides
  • A method for in situ culture of amniocytes and karyotype processing and analysis on glass slides

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Example 1: Synchronous culture of amniocytes on domestic slides and imported slides

[0031] (1) Inoculation: Select 10 amniotic fluid samples, each sample takes 18-22ml. Aseptically divide into 2 conical centrifuge tubes, 9-11ml / tube, balance and centrifuge for 10min at 1200rpm, keep 0.5ml of cell suspension (amniotic fluid cell pellet + a little amniotic fluid) in each tube, add 1.5ml of amniotic fluid culture medium to each tube , respectively tiled on the cell-adherent surface of the domestic glass slide and the imported original glass slide culture flask ( figure 2 b) at 37°C, 5% CO 2 Incubate for 2 days under conditions with minimal mechanical disturbance, and after 2 days at the mark on the slide ( figure 2 a) Add another 3ml of amniotic fluid medium, mix the newly added medium with the old medium, and place it again at 37°C, 5% CO 2 conditions for 5 days.

[0032] (2) Change the medium: After the amniotic fluid cells have been cultured for 7-8 days, place ...

Embodiment 2

[0047] Example 2: Effects of different hypotonic fluids on cleavage phase during the harvesting of amniotic fluid cultured on domestic slides

[0048] 1) Inoculation: Take 24 amniotic fluid samples, 5ml for each sample, mix them evenly (to eliminate the differences among the samples), divide into 12 bottles, 2 bottles for each condition. 10ml / tube, balance and centrifuge for 10min at 1200rpm, take 0.5ml of cell suspension (amniotic fluid cell pellet + a little amniotic fluid) from each tube, add 1.5ml of amniotic fluid culture medium to each tube, and spread them on domestic glass slides and imported original loading The cell-attached side of the slide culture flask ( figure 2 b) at 37°C, 5% CO 2 Incubate for 2 days under the conditions of minimal mechanical disturbance. After 2 days, mark the slide ( figure 2 a) Add another 3ml of amniotic fluid medium, mix the newly added medium with the old medium, and place it again at 37°C, 5% CO 2 conditions for 5 days.

[0049] 2)...

Embodiment 3

[0055] Example 3: Effects of Different Pre-fixed Solutions on Split Phases During Amniotic Fluid Harvesting in Domestic Slide Culture

[0056] 1) Inoculation: Take 20 amniotic fluid samples, 5ml for each sample, mix them evenly (to eliminate the differences among the samples), divide into 10 bottles, 2 bottles for each condition. 10ml / tube, balance and centrifuge for 10min at 1200rpm, take 0.5ml of cell suspension (amniotic fluid cell pellet + a little amniotic fluid) from each tube, add 1.5ml of amniotic fluid culture medium to each tube, and spread them on domestic glass slides and imported original loading The cell-attached side of the slide culture flask ( figure 2 b) at 37°C, 5% CO 2 Incubate for 2 days under conditions with minimal mechanical disturbance, and after 2 days at the mark on the slide ( figure 2 a) Add another 3ml of amniotic fluid medium, mix the newly added medium with the old medium, and place it again at 37°C, 5% CO 2 conditions for 5 days.

[0057]...

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Abstract

The present invention provides a method for in situ culture of amniotic fluid cells and karyotype processing and analysis on glass slides. The present invention is further improved on the basis of previous research, and uses specific concentrations and types of hypotonic fluid and pre-fixed fluid to strictly control chromosomes. When dispersing, the temperature and humidity of the environment are dispersed to obtain abundant high-quality split phases. This technology will provide a basic technology platform for the high-throughput automatic scanning and capture system, establish an automated film reading process, and improve clinical work efficiency.

Description

technical field [0001] The invention relates to the field of amniotic fluid cell culture, in particular to a method for in-situ culture of amniotic fluid cells on glass slides and karyotype processing and analysis. Background technique [0002] Amniotic fluid cell culture and karyotype analysis are the key technologies for prenatal diagnosis. This technology needs to go through amniotic fluid cell adhesion, harvesting (hypoosmotic, pre-fixed), film production (chromosomal dispersion), karyotype taking under a microscope and karyotype analysis. And many other steps to complete the final clinical report. With the continuous popularization of prenatal screening and the rapid increase in the demand for prenatal diagnosis, traditional amniotic fluid cell culture (time-consuming and labor-intensive) cannot meet clinical needs, and the traditional amniotic fluid cell culture has many steps and unstable production quality , not suitable for quality control of prenatal diagnosis. I...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/071C12Q1/68
Inventor 唐少华林小玲郑义
Owner WENZHOU CENT HOSPITAL
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