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A method for detecting cytotoxicity of drug to be tested on target cells and special cell chip thereof

A cell chip and cell technology, applied in the detection of programmed cell death, methods of supporting/immobilizing microorganisms, biochemical equipment and methods, etc., can solve the problem of high cost of large-scale instruments, cumbersome operation, and inability to truly reflect the physiological state of individual cells, etc. question

Active Publication Date: 2018-08-28
TSINGHUA UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Traditional biochemical analysis with cells as the object uses a large number of cells to obtain the overall mean value of relevant parameters through large instruments to describe the physiological activity of cells, but analysis methods based on cell populations often cannot truly reflect the physiological state of individual cells , and the cost of large-scale instruments is high, the operation is cumbersome, and special technical personnel are required for maintenance, which has certain limitations in practical applications

Method used

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  • A method for detecting cytotoxicity of drug to be tested on target cells and special cell chip thereof
  • A method for detecting cytotoxicity of drug to be tested on target cells and special cell chip thereof
  • A method for detecting cytotoxicity of drug to be tested on target cells and special cell chip thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0100] Embodiment 1, the making of SF767 cell chip

[0101] The process of making SF767 cell chip is detailed in figure 1 .

[0102] 1. Make a mask: Use AutoCAD2010 drawing software to design and manufacture the graphics of the SF767 cell chip, convert the design graphics into graphics files, and use a high-resolution printer (≥1200dip) to print the graphics files on the chrome plate glass. The chrome plate glass is for the mask.

[0103] 2. Photolithography: After completing step 1, use a mixed solution of concentrated sulfuric acid and hydrogen peroxide (obtained by mixing concentrated sulfuric acid and hydrogen peroxide at a volume ratio of 3:1), acetone and ethanol to clean the silicon wafer, and dry it with an air gun , and then apply SU-8 2050 negative photoresist on the silicon wafer, and then pre-bake (the effect is to fully volatilize the solvent in the SU-8 2050 negative photoresist, and the dried colloid can increase the contact with the silicon wafer. Adhesion a...

Embodiment 2

[0114] Example 2, application of SF767 cell chip to detect cytotoxicity and gene damage of the drug to be tested

[0115] The flow chart of the application of SF767 cell chip to detect the cytotoxicity and gene damage of the drug to be tested is as follows image 3 shown.

[0116] The drug to be tested is nimustine (ACNU), an alkylating agent-type antineoplastic drug, which can cause cross-linking between DNA strands, block DNA replication and transcription, and induce cell apoptosis. The experiment was repeated three times, and the steps for each repetition were as follows:

[0117] 1. Dissolve and dilute nimustine with MEM resistant medium, prepare 300mg / mL nimustine stock solution, filter and sterilize through a 0.22μm filter membrane, and store at -20°C. When in use, dilute the nimustine mother solution with PBS buffer solution of pH 7.4 and 0.01M to obtain a working solution 1 with a nimustine concentration of 0.05mM, a working solution 2 with a nimustine concentration ...

Embodiment 3

[0122] Example 3. Application of SF767 cell chip immunofluorescence in situ detection of the effect of the drug to be tested on the expression of the target protein in the cell

[0123] The flow chart of using SF767 cell chip immunofluorescence to detect the effect of the drug to be tested on the expression of the target protein in the cell is as follows Figure 5shown. The drug to be tested is nimustine, an alkylating agent antitumor drug, which can lead to cross-linking between DNA strands, block DNA replication and transcription, and induce cell apoptosis. The experiment was repeated three times, and the steps for each repetition were as follows:

[0124] 1. Dissolve and dilute nimustine with MEM resistant medium, prepare 300mg / mL nimustine stock solution, filter and sterilize through a 0.22μm filter membrane, and store at -20°C. When in use, dilute the nimustine mother solution with PBS buffer solution of pH 7.4 and 0.01M to obtain a working solution 1 with a nimustine c...

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Abstract

The invention discloses a method for detecting the cytotoxicity of a to-be-tested medicine to a target cell and a cell chip specially used by the method. The cell chip provided by the invention comprises a supporting medium and a sepharose gel film attached to the upper part of the supporting medium, wherein a plurality of microtraps are distributed in the sepharose gel film, and the size of each microtrap is matched with that of the individual target cell; and the individual target cell is attached to the inside of each microtrap. The experiment proves that the cell chip provided by the invention can detect the cytotoxicity of the to-be-tested medicine to the cell and / or detect the gene damage of the to-be-tested medicine to the cell and / or detect the influences of the to-be-tested medicine to the expression of interest protein in the cell and / or screen high-content drugs having effects for the cells. The cell chip provided by the invention has important application values.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a method for detecting the cytotoxicity of a drug to be tested on target cells and a special cell chip thereof. Background technique [0002] With the development of modern science and technology, the technology in the fields of computer simulation design, chemical synthesis, biological extraction and natural product extraction has advanced by leaps and bounds, making the acquisition of target compounds faster and more efficient. The number of samples in the compound library ranges from hundreds to millions, and rapid screening of pharmacologically active compounds or lead compounds is one of the research hotspots in the pharmaceutical industry at present. High-content drug screening refers to the simultaneous detection of the effects of the drug to be tested on cell morphology, growth, differentiation, migration, apoptosis, metabolic pathways and signal transduction on t...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12M3/00C12M1/34C12Q1/02
CPCC12M23/12C12M25/00C12N2503/02G01N33/5008G01N33/5014G01N2500/10
Inventor 梁琼麟李莉莉
Owner TSINGHUA UNIV
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