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Carrier for in-situ analysis of intracellular protein compound as well as preparation method and application of carrier

A technology for in situ analysis of protein complexes, applied in the field of carrier for in situ analysis of intracellular protein complexes and its preparation and application, can solve the difficulties of multiple side reactions and in situ targeted analysis of intracellular protein complexes , strong activity of cross-linking agent and other issues, to achieve low cytotoxicity, solve the difficulty of in situ targeted analysis, and solve the effect of strong activity

Active Publication Date: 2020-12-15
DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
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  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0008] The purpose of the present invention is to provide a carrier for in situ analysis of intracellular protein complexes and its preparation method and application, through the carrier of the present invention for in situ analysis of intracellular protein complexes effectively solves the cross-linking agent activity Strong, many side effects, in addition to solving the problem of in situ targeted analysis of intracellular protein complexes

Method used

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  • Carrier for in-situ analysis of intracellular protein compound as well as preparation method and application of carrier

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Experimental program
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Effect test

Embodiment 1

[0029] With 2 mass parts of castor oil polyoxyethylene ether, molecular weight is 15 mass parts of polylactide-glycolide of 10000, 1 mass part of didodecyl dimethyl ammonium bromide, crosslinking agent (disuccinimide Suberate) 4 mass parts are dissolved in the dichloromethane of 500 mass parts. The above mixture was poured into a 1% by mass polyvinyl alcohol aqueous solution, and ultrasonicated for 2 minutes, and the carrier was solidified with a 1% by mass polyvinyl alcohol aqueous solution after the ultrasonication was completed. The TEM photo of the carrier obtained in this embodiment is as follows figure 1 shown. The particle size and surface potential of the prepared materials are shown in Table 1.

[0030] Table 1. DLS test data (1, 2, 3 are three DLS tests for the carrier)

[0031] 1 2 3 Particle size / nm 274.6 268.8 273.7 PDI 0.237 0.181 0.135 Zeta potential / mv 38 39.9 41

[0032] Depend on figure 1 It can be seen from the T...

Embodiment 2

[0035]With castor oil polyoxyethylene ether 3 mass parts, molecular weight is 15 mass parts of polylactide-glycolide of 20000, didodecyl dimethyl ammonium bromide 1 mass part, crosslinking agent (disuccinimide Suberate) 5 mass parts are dissolved in the dichloromethane of 400 mass parts. The above mixture was poured into 1% polyvinyl alcohol aqueous solution, ultrasonicated, and the carrier was further solidified with 1% polyvinyl alcohol solution. The particle size of the prepared carrier is 126.6nm±0.75nm, and the potential (Z-potential: 46.6±0.379mv) meets the conditions for it to be used as a transmembrane transporter.

[0036] In the study of the in situ analysis of the cross-linking agent carrier for protein complexes, the carrier and the cells were co-incubated in 1640 culture medium. After incubation for 3 hours, the cells were collected, and then the proteins and protein complexes were extracted from the cells using ionic liquids. The extracted protein and protein co...

Embodiment 3

[0038] With 5 mass parts of castor oil polyoxyethylene ether, molecular weight is 15 mass parts of polylactide-glycolide of 80000, 2 mass parts of didodecyl dimethyl ammonium bromide, crosslinking agent (disuccinimide Suberate) 8 mass parts are dissolved in the dichloromethane of 700 mass parts. The above mixture was poured into 0.5% polyvinyl alcohol aqueous solution, ultrasonicated, and the carrier was further solidified with 2% polyvinyl alcohol solution. The particle size of the prepared carrier is 236.6nm±0.92nm, and the potential (Z-potential: 49±0.379mv) meets the conditions for it to be used as a transmembrane transporter.

[0039] In the study of the cross-linking agent carrier used in the in situ analysis of protein complexes, the carrier and cells were co-incubated in 1640 culture medium. After incubation for 6 hours, the cells were collected, and then the proteins and protein complexes were extracted from the cells using ionic liquids. The extracted protein and pr...

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Abstract

The invention relates to a carrier for in-situ analysis of an intracellular protein compound as well as a preparation method and application of the carrier. The carrier for in-situ analysis of the intracellular protein complex is used for conveying a chemical cross-linking agent for protein complex analysis to a specific part in a cell in a transmembrane delivery manner and releasing the chemicalcross-linking agent, so that the protein complex is subjected to in-situ cross-linking. And analyzing the spatial-temporal dynamic change rule of the intracellular protein compound by combining a massspectrometry technology. The method has great significance in understanding the life process, revealing the disease mechanism, screening biomarkers and searching drug targets.

Description

technical field [0001] The invention relates to a carrier for in-situ analysis of intracellular protein complexes, a preparation method and application thereof. The invention belongs to functional biological material and its preparation and application in in situ analysis of protein complex. Background technique [0002] Protein complexes are important building blocks of protein machinery. High-dimensional signal transduction networks dynamically assembled by protein complexes run through different subcellular organelles and regulate almost all life processes. Analyzing the temporal and spatial dynamic changes of protein complexes in complex systems and drawing interaction networks are of great significance for understanding life processes, revealing disease mechanisms, screening biomarkers, and finding drug targets. [0003] Chemical cross-linking combined with mass spectrometry (referred to as cross-linking mass spectrometry or CXMS) is a new method developed in recent y...

Claims

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Application Information

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IPC IPC(8): G01N30/02G01N30/06
CPCG01N30/02G01N30/06G01N2030/067
Inventor 张丽华陈玉宛杨开广周雯裴家琪梁振张玉奎
Owner DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
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