Method for capturing cells or biomolecules in solutions

A technology of biomolecules and cells, applied in the methods of supporting/immobilizing microorganisms, cell dissociation methods, biochemical equipment and methods, etc., which can solve the problems of low specificity, inability to collect and re-cultivate circulating tumor cells, and rely on old-fashioned chips.

Active Publication Date: 2017-11-10
苏州博福生物医药科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Generally, there are only 1-10 circulating tumor cells in 10ml blood of cancer patients, and their slow capture speed or poor specificity is an urgent problem to be solved for rapid detection of patient blood samples
[0003] The current sorting methods such as the traditional immunomagnetic bead sorting method have good repeatability, high sensitivity, good specificity, and can quantitatively analyze circulating tumor cells, but the operation speed is slow and the throughput is small
Although membrane microfiltration technology and gradient density centrifugation method are simple to operate and have good cell viability after separation, they have low specificity and high false positive rate
Microfluidic technology is simple to operate and requires a small amount of antibodies, but it is costly and has a high false negative rate
Moreover, microfluidic technology companies are now mainly targeting scientific research customers, who need to mix multiple reagents in advance, rely too much on old-fashioned chips, and have difficulties in industrialization, so it is difficult to enter the field of in vitro diagnostics
Others, such as the Cell Search system in the United States, have high sensitivity and high specificity, but consume a lot of blood, require a large amount of antibodies, and are expensive, and cannot capture living cells and collect and re-culture circulating tumor cells, so they can only do DNA sequencing cannot be used for RNA sequencing, and cannot be used as a medication guide

Method used

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  • Method for capturing cells or biomolecules in solutions
  • Method for capturing cells or biomolecules in solutions
  • Method for capturing cells or biomolecules in solutions

Examples

Experimental program
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Effect test

example 1

[0080] Herein, a preparation method of the capture sieve for sorting CTCs is presented. The preparation method includes:

[0081] (I) selection of sieves;

[0082] Gold sieve: Choose a gold sieve with pores (eg, 64 µm or 40 µm) to maximize contact time with tumor cells while preventing the risk of clogging. The size of the gold sieve used in the following tests is 2×2mm 2 . Larger sizes are preferred depending on the specific operation.

[0083] Stainless steel and gold sieves: 51µm pores are selected and AuPd is coated by magnetron sputtering. Such sieves are cheaper and mechanically stronger than gold, so they are more suitable for integration in devices.

[0084] (II) Pre-functionalization;

[0085] Various cleaning methods were used to prepare the sieves prior to functionalization, including autoclaving, oxygen plasma cleaning, and ultrasonic cleaning in various solutions, including piranha solutions. For example, the best results for a 64 µm gold sieve included son...

example 2

[0095] Experiments were performed to demonstrate the efficiency of the screen in capturing EpCAM expressed by tumor cells. In this case, the mesh opening is 51 μm.

[0096] Cell Selection - Cells with high expression levels of EpCAM protein (such as CaCo2 and MCF7 cells) were used.

[0097] Cell Growth - Grow in DMEM buffer at 37°C.

[0098] Cell preparation and incubation of screens - CaCo2 and MCF7 cells differentiated 1:2. The test was performed on a rotating 37°C hot plate. After isolation, cells were diluted 1:10 in DMEM buffer, and 0.5 ml was used to incubate the capture screen for 1 hour.

[0099] For washing of non-specifically bound cells, the sieves were rinsed with pure water, incubated in pure aqueous solution for 2 minutes, and then rinsed again before microscopic observation.

[0100] Microscopic evaluation of trapped cells: photograph of the sieve taken by the microscope ( Figure 5 a and 5b). Figure 5 a shows CaCo2 cells expressing the capture molecule o...

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Abstract

The invention discloses a method for capturing cells or biomolecules. The method comprises the following steps: (I) enabling a medium containing target cells or biomolecules to enter a device comprising a capturing mechanism for capturing the target cells or biomolecules; (II) enabling the medium to flow through the capturing mechanism so that the target cells or biomolecules are bound to the capturing structure; (III) removing the needless impurities, cells or molecules which are not specifically bound to the capturing mechanism; (IV) enabling the target cells or biomolecules to be separated from the capturing mechanism; and (V) collecting the target cells or biomolecules. The capturing mechanism comprises one or more stacked capturing sieves, wherein each capturing sieve comprises a net-shaped base body and a capturing layer formed on the net-shaped base body, and the capturing layer contains capturing matter which can be in specific binding with the target cells or biomolecules. The method disclosed by the invention has high specificity and high flux, is suitable for capturing the cells or capturing the biomolecules in the solutions by adopting molecules expressed by cells, and is particularly suitable for capturing and sorting circulating tumor cells.

Description

technical field [0001] The present invention relates to a device for capturing a cell or capturing a biomolecule in solution by a biomolecule expressed by the cell, and more particularly to a device for capturing a target cell (eg, circulating tumor cells) or A capture screen for capturing target biomolecules in a solution and a device having such a capture screen. Background technique [0002] Circulating tumor cells, that is, tumor cells circulating in the blood, are considered to have a significant relationship with distant metastasis of tumors and other issues. Generally, there are only 1-10 circulating tumor cells in 10ml blood of cancer patients, and their slow capture speed or poor specificity is an urgent problem to be solved for rapid detection of patient blood samples. [0003] Current sorting methods such as the traditional immunomagnetic bead sorting method have good repeatability, high sensitivity, and good specificity, and can quantitatively analyze circulatin...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12M1/12C12M1/34C12N5/09
CPCC12M41/36C12M47/04C12N5/0693C12N2509/00
Inventor 查梅特·杰罗姆颜菁鲍尔·沃尔夫冈安德烈斯·克里斯蒂安余子夷杨洋
Owner 苏州博福生物医药科技有限公司
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