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Cell membrane chromatographic column for identifying microscale target component in complex sample and preparation method thereof, and identification method of target component

A complex sample and cell membrane technology, applied in the field of cell membrane chromatography, can solve the problems of long analysis time, poor resolution, low efficiency, etc., and achieve the effect of improving analysis efficiency, improving screening efficiency and simple steps.

Inactive Publication Date: 2017-04-19
XI AN JIAOTONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] (1) The efficiency is low, and the "target component" cannot be effectively determined;
[0004] (2) The analysis time is long, there are many complex sample components, the chromatographic analysis time is generally more than 60min, and the organic solvent consumption is relatively large;
[0005] (3) The resolution is poor, and it is difficult to achieve baseline separation of components in complex samples, and the trace "target components" are easily "buried" by the constant components and cannot be measured

Method used

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  • Cell membrane chromatographic column for identifying microscale target component in complex sample and preparation method thereof, and identification method of target component
  • Cell membrane chromatographic column for identifying microscale target component in complex sample and preparation method thereof, and identification method of target component

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preparation example Construction

[0031] The preparation method of the cell membrane chromatographic column for identifying trace target components in complex samples provided by the present invention comprises the following steps:

[0032]1) Take the cell suspension with high expression of specific receptors, centrifuge in a high-speed centrifuge at 1000rpm and 4°C for 5min, pour off the supernatant, and take the bottom cells; wash the centrifuged cells twice with 5mL normal saline , pipet evenly into a 10mLEP tube, and centrifuge twice at 1000rpm at 4°C; then add 5mL Tris-Hcl to the centrifuged cells, and ultrasonically break them for 30min in an ice bath, then use a cell disruptor (400W , 8 times, 3s each time) After breaking, centrifuge at 5000rpm for 10min, remove the organelles, and take the supernatant; balance the supernatant, centrifuge at 12000g for 20min, after centrifugation, stick the membrane on the wall of the EP tube, add After 3mL of normal saline is blown away, add normal saline to 10mL; obta...

Embodiment 1

[0040] 1. Culture of HMC-1 cells

[0041] 1) Recovery of cells

[0042] Take out the pre-frozen tube containing HMC-1 cells from the liquid nitrogen tank, quickly put it into a water bath that has been preheated to 37°C, and shake it gently to melt the frozen HMC-1 cells as soon as possible . After the cells are thawed, take out the cryopreservation tube, disinfect it with 75% alcohol and open it in an ultra-clean bench, pipette the HMC-1 cell suspension into a 10mL centrifuge tube, add 10 times more culture medium dropwise, and mix well by pipetting Afterwards, centrifuge at 1000rpm at low speed for 5 minutes, remove the supernatant, add a small amount of culture medium to the cell pellet, blow and mix, transfer to a cell culture bottle, add appropriate culture medium to dilute, and place the culture bottle in a cell culture incubator for static culture. The medium was changed daily to continue culturing.

[0043] 2) Subculture of cells

[0044] When the resuscitated HMC-...

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Abstract

The invention discloses a cell membrane chromatographic column for identifying a microscale target component in a complex sample and a preparation method thereof, and an identification method of the target component. The cell membrane chromatographic column (identification column for short) for identifying the microscale target component in the complex sample is filled with a cell membrane chromatographic stationary phase for expression of specific receptors. Screening specificity of the target component in the complex sample can be enhanced, and it is of great significance for enhancing screening efficiency and expounding mechanism of action. According to the identification method, by specific identification of the target component in the complex sample through the identification column and by online enrichment of the microscale target component through an enriching column, separation and identification are further carried out by high performance liquid chromatography-mass spectrometry. The method of the invention has the characteristics of efficient identification and online enrichment of the microscale target component in the complex sample, can help enhance analysis efficiency, raise sensitivity for analysis and realize high-efficiency screening of the target component in the complex sample.

Description

technical field [0001] The invention belongs to the technical field of cell membrane chromatography, and relates to a cell membrane chromatographic column for identifying and identifying trace "target components" in complex samples and a method for identifying and identifying trace "target components" in complex samples using the cell membrane chromatographic column. Background technique [0002] Complex samples (such as traditional Chinese medicine extracts and their preparations) have complex components (hundreds of species), and high-throughput screening is an important method for discovering target components in complex systems. Many high-throughput screening methods have been reported, such as direct Screening based on receptors or enzymes, virtual screening for computer-aided drug design, etc. It is difficult to identify all the components in complex samples by traditional high-performance liquid chromatography analysis method for fingerprint analysis. with the follow...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N30/60B01J20/282
CPCG01N30/6052B01J20/282
Inventor 贺浪冲孙萌林园园韩省力马维娜吕艳妮杨柳
Owner XI AN JIAOTONG UNIV
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