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Fluorescent tracing system and method suitable for researching in-vitro and in-vivo distribution of lipidosome

A fluorescent tracking, liposome technology, applied in the directions of liposome delivery, preparations for in vivo experiments, fluorescence/phosphorescence, etc., can solve problems such as the difficulty of studying the distribution of liposomes in vivo and in vitro, and avoid intra-group differences. , the purification method is simple, and the effect is easy to discharge

Active Publication Date: 2020-04-03
SHANGHAI JIAO TONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] In view of the shortcomings of the prior art described above, the purpose of the present invention is to provide a fluorescent tracer system and method suitable for studying the distribution of liposomes in vivo and in vitro, which is used to solve the difficulties in the study of liposome distribution in vivo and in vitro The problem

Method used

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  • Fluorescent tracing system and method suitable for researching in-vitro and in-vivo distribution of lipidosome
  • Fluorescent tracing system and method suitable for researching in-vitro and in-vivo distribution of lipidosome
  • Fluorescent tracing system and method suitable for researching in-vitro and in-vivo distribution of lipidosome

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0081] Preparation and characterization of liposomes labeled with trans-cyclooctene in Example 1

[0082] (1) trans cyclooctene active ester (TCO-NHS) and amino lipid (in this example, DSPE-PEG2000-NH 2 For example) were dissolved in organic solvents (such as dichloromethane, chloroform, absolute ethanol, etc.), so that the final concentration of the solution was 10mg / mL; the trans-cyclooctene active ester solution was added to the amino lipid solution , so that the molar number of trans-cyclooctene active ester is 2 to 5 times that of the amino lipid, add triethylamine with a volume ratio of 0.3% to 0.5%, shake gently at room temperature, react for 8 hours, and use ethyl acetate The unreacted trans-cyclooctene active ester is removed by washing with a mixed solution of ether to obtain trans-cyclooctene lipid (DSPE-PEG2000-TCO). The organic solvent in the lipid solution of trans-cyclooctene is removed, and an appropriate amount of deionized water is added to make the concentr...

Embodiment 2

[0089] Embodiment 2 trans-cyclooctene liposome preparation stability research

[0090] The trans-cyclooctene liposomes prepared in Example 1 (the molar ratio of trans-cyclooctene in lipids is 1%, 3%, 5%) were redispersed in deionized water, PBS, HEPES, 1640 culture medium and saline. At different time points, the particle size, polydispersity index (PDI) and zeta potential of the particle size were measured using a dynamic light scattering nanometer particle size analyzer (DLS) to investigate its stability.

[0091] Drug-loaded trans-cyclooctene liposomes (the molar ratio of trans-cyclooctene is 1%, 3%, and 5%) were analyzed by detecting ultraviolet absorption (UV) with a microplate reader after dialysis to analyze drug leakage at different time points In this case, doxorubicin was used as the model drug.

[0092] Such as Figures 5a-5dAs shown, the particle size of trans-cyclooctene liposomes (1%) has no significant change in deionized water, PBS, HEPES, 1640 medium and ph...

Embodiment 3 4

[0094] The synthesis of embodiment 3 tetrazine fluorescent probes

[0095] Tetrazine active ester (Tetrazine-PEG5-NHS Ester) and near-infrared fluorescent dye (using Cy5.5 in this example) were dissolved in organic solvent respectively, and the two were mixed so that the molar number of tetrazine active ester was near-infrared fluorescent dye 3 to 5 times that of fluorescent dyes, adding triethylamine with a volume ratio of 0.3% to 0.5%, reacting at room temperature with gentle shaking for 12 hours in the dark, and washing with ether and ethyl acetate after the reaction to separate and obtain blue Solid, dark blue powder obtained after freeze-drying, which is the tetrazine near-infrared fluorescent probe (Tz-Cy5.5). Before use, it can be redissolved in buffers such as PBS, normal saline, and serum-free medium.

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Abstract

The invention discloses a fluorescent tracing system and method suitable for researching in-vitro and in-vivo distribution of lipidosome, and the method is based on a biological orthogonal reaction principle and is combined with fluorescent tracing technology. The system mainly comprises a trans-cyclooctene liposome and a tetrazine fluorescent probe. By adopting the method, the in-vitro and in-vivo distribution of the drug-loaded liposome can be researched, and a convenient and feasible method is provided for researching the in-vivo distribution of the liposome.

Description

technical field [0001] The invention relates to the field of drug research in vivo, in particular to a fluorescent tracer system and method suitable for studying the distribution of liposomes in vivo and in vitro. Background technique [0002] Liposome is a spherical phospholipid bilayer vesicle made artificially, and its lumen can contain drugs. Because liposomes have similar components and structures to biological membranes, they have good biocompatibility, and at the same time have the advantages of biodegradability, no obvious toxicity, and low immunogenicity. Since there is a certain correlation between the distribution of liposome vesicles and drugs in vivo before drug release, the method of studying the distribution of liposome vesicles can be used to study the distribution of liposome drugs in vivo. At present, the research on the distribution of liposomes in the body generally relies on the method of detecting the drug content in the tissue. This method has the fol...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/64G01N15/14A61K49/00
CPCG01N21/6428G01N21/6402G01N21/6458G01N21/6486G01N15/14A61K49/0021A61K49/0084G01N2015/1006
Inventor 彭金良陈月潭陈阳徐宇虹
Owner SHANGHAI JIAO TONG UNIV
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