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Chlorine imaging method of central nervous system based on laser confocal scanning microscope system

A laser confocal and scanning microscope technology, applied in fluorescence/phosphorescence, material excitation analysis, preparation of test samples, etc., to achieve reliable results, less damage, and less autofluorescence

Active Publication Date: 2010-07-28
FOURTH MILITARY MEDICAL UNIVERSITY
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AI Technical Summary

Problems solved by technology

However, how to measure the concentration of chloride ions in specific cells (mainly neurons) in the nervous system, how to visualize the concentration of chloride ions in cells, and how to monitor the changes of chloride ion concentration in real time and dynamically have not been reported in China.

Method used

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Experimental program
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Embodiment Construction

[0014] The chlorine imaging method of the central nervous system based on the laser confocal scanning microscope system follows the following steps: (1) co-incubation of the brain slices with the chloride ion fluorescent probe MQAE; (2) imaging and recording with the laser confocal microscope; (3) calibration of the cells MQAE fluorescence value of the internal chloride ion fluorescent probe.

[0015] 1. Co-incubation of brain slices with chloride ion fluorescent probe MQAE:

[0016] (1) A brain slice with a thickness of 200-300 μm was placed flat on a circular cover glass pre-placed in a 24-well plate for cell culture. Circular coverslips were coated with a polytyrosine solution at a concentration of 1 mg / ml before use to facilitate cell attachment.

[0017] (2) Use a neutral buffer solution fed with a mixed gas, called Krebs-HEPES buffer solution, to wash to remove impurities. It is required to wash 3 times, each time for 5 minutes.

[0018] (3) Add a small amount (100-200...

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Abstract

The invention discloses a chlorine imaging method of a central nervous system based on a laser confocal scanning microscope system, comprising the following steps of: (1) together incubating a brain slice and chloridion fluorescence probe N-(ethoxycarbonylmethyl)-6-methoxyquinolinium bromide (MQAE); (2) imaging and recording by using a laser confocal microscope; and (3) correcting the fluorescent value of the MQAE in cells. The invention mainly has the advantages of quantitatively detecting the chlorine ion concentrations in nerve cells and change conditions thereof and visualizing the chlorine ion concentrations in a fluorescent form for the detected nerve cells, and having small self-emitting fluorescence and little damage on the cells because the adopted MQAE is excited in visible light.

Description

Technical field: [0001] The invention belongs to the field of laser confocal imaging, and relates to a central nervous system chlorine imaging method based on a laser confocal scanning microscope system. Background technique: [0002] Chloride ion is the most abundant anion in the body. Chloride channels that regulate the concentration of chloride ions inside and outside cells are widely present in the cell membrane and organelle membrane of the body. Chloride channels play an important role in various activities and regulatory processes of cells, such as cell proliferation, apoptosis, regulation of cell excitability, pH regulation, volume regulation and immune response. Therefore, the homeostasis of chloride ions is a necessary condition for maintaining various metabolic activities of cells. Once the homeostasis of chloride ions is disrupted, cells will suffer functional or organic damage or even die. [0003] The laser confocal scanning microscope system is an image dete...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/64G01N1/28
Inventor 王亚云杨雁灵季茹李俊杰千年松武胜昔李云庆魏燕燕
Owner FOURTH MILITARY MEDICAL UNIVERSITY
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