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Method for acquiring single cell array

An acquisition method, single-cell technology, applied in the field of acquisition of single-cell arrays, can solve problems such as inability to fit tightly, easy to slide left and right, easy deformation of tweezers, etc., to achieve fast drilling speed, easy to obtain raw materials, and simple and easy factory operation Achieved effect

Active Publication Date: 2021-02-05
LINYI UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Through-hole chips reported in the past are generally prepared by using SU8 or PDMS (polydimethylsiloxane) materials through micro-nano processing methods, and the thickness is generally 20-30 μm. There are three types of through-hole chips with this thickness. There are two problems: ① After sticking to the hard substrate, it cannot fit tightly, and it is easy to slide left and right, so bonding or clamps are generally required to fix it; ② This thickness is equivalent to the size of a cell. Due to fluid resistance, a single Cells cannot enter the wells through free settling, so external forces such as centrifugation for a few minutes are often required to drive cells into deep wells; ③ SU8 through-hole chips are very brittle and easily broken under external force, followed by SU8 chips after bonding It is difficult to peel off the film, and PDMS is too soft, and it is easy to deform and stick when picked up by tweezers

Method used

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  • Method for acquiring single cell array
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  • Method for acquiring single cell array

Examples

Experimental program
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Effect test

Embodiment 1

[0052] The method for obtaining a single-cell array (substrate+ultra-thin through-hole metal sheet), specifically includes the following steps:

[0053] (1) Laser drilling is used to prepare ultra-thin through-hole metal sheets with a thickness of 5 μm, a hole diameter of 10 μm, and a hole spacing of 80 μm, and the through-hole area is cleaned for standby; the specific operation of cleaning is: firstly, the ultra-thin through-hole metal sheet The through-hole metal sheet is placed in absolute ethanol for ultrasonic treatment with a power of 0.3W / cm 2 , the temperature is 20°C, the time is 5min, then use absolute ethanol and deionized water to clean in turn, then blow dry with nitrogen, and sterilize under high pressure. Finally, use a microscope to observe whether the through hole area is completely cleaned, otherwise continue according to the above steps cleaning;

[0054] (2) Attach the ultra-thin through-hole metal sheet to the culture dish and press it tightly until there...

Embodiment 2

[0058] The method for obtaining a single-cell array (substrate+buffer layer+ultra-thin through-hole metal sheet), specifically includes the following steps:

[0059] (1) Micromachining is used to prepare an ultra-thin through-hole metal sheet with a thickness of 5 μm, a hole diameter of 25 μm, and a hole spacing of 80 μm, and the through-hole area is cleaned for standby; wherein, the specific operation of cleaning is: firstly, the ultra-thin through-hole metal sheet The perforated metal sheet is placed in absolute ethanol for ultrasonic treatment with a power of 0.3W / cm 2 , the temperature is 30°C, the time is 8min, then use absolute ethanol and deionized water to clean in sequence, then blow dry with nitrogen, and sterilize under high pressure. Finally, use a microscope to observe whether the through hole area is completely cleaned, otherwise continue according to the above steps cleaning;

[0060] (2) Spread a layer of polydimethylsiloxane buffer layer on the petri dish fir...

Embodiment 3

[0064] The method for obtaining a single-cell array (substrate+coating layer+ultra-thin through-hole metal sheet), specifically includes the following steps:

[0065] (1) Laser drilling is used to prepare ultra-thin through-hole metal sheets with a thickness of 5 μm, a hole diameter of 25 μm, and a hole spacing of 100 μm, and the through-hole area is cleaned for standby; the specific operation of cleaning is: first The through-hole metal sheet is placed in absolute ethanol for ultrasonic treatment with a power of 0.4W / cm 2 , the temperature is 40°C, the time is 10min, then use absolute ethanol and deionized water to clean in sequence, then blow dry with nitrogen, sterilize with ultraviolet light, and finally use a microscope to observe whether the through hole area is completely cleaned, otherwise continue according to the above steps cleaning;

[0066] (2) Use a pipette gun to pipette the melted fibronectin into the petri dish and cover it completely, then draw out the remai...

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Abstract

The invention discloses a method for acquiring a single cell array, wherein the method specifically comprises the following steps: (1) preparing an ultrathin through hole metal sheet, and cleaning forlater use; (2) tightly attaching the ultrathin through hole metal sheet to a substrate, and pressing the ultrathin through hole metal sheet until no bubble exists; (3) extracting a cell suspension, placing the cell suspension in a through hole area of the ultrathin through hole metal sheet, and standing in an incubator; and (4) pumping out the cell suspension, washing the through hole area with abuffer solution, and then taking down the ultrathin through hole metal sheet to obtain the single cell array. The method does not need bonding or fixture fixation, does not need a sample pool or centrifugation, is simple in operation and high in efficiency, and provides great help for large-area acquisition of the single cell array; the ultrathin through hole metal sheet is wide in raw material source, easy to obtain, low in processing cost, capable of being subjected to ultrasonic, organic, boiling, drying and other cleaning steps and capable of being repeatedly used, and thus the cost is reduced.

Description

technical field [0001] The invention relates to the technical field of biochips, and more specifically relates to a method for obtaining a single cell array. Background technique [0002] The data obtained by traditional cell detection methods are based on the average value of a large number of cells, and the specific information of a single cell obtained by traditional cell detection methods is often considered "wrong" or "noise" and ignored or discarded. In modern biomedical research, especially in tumor research, due to the heterogeneity of cells, traditional cell detection methods often cannot meet the requirements for accurate analysis of cell signals. [0003] Single-Cell Analysis (Single-Cell Analysis) is an analysis method that focuses on the unique response of a single cell to different stimulus signals. This analysis method overcomes the shortcomings of traditional cell detection methods. Diagnostics and personalized medicine are of great significance. [0004] S...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): B01L3/00C40B40/02
CPCB01L3/502707B01L3/502753C40B40/02
Inventor 孙英男宋宇涵田晴晴刘建红郭文婷张书圣
Owner LINYI UNIVERSITY