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Method for recording Nav 1.5 sodium channel current by separating cavy ventricular muscle cells

A technique for ventricular myocytes and sodium channels, which is applied in the field of separating guinea pig ventricular myocytes and recording Nav1.5 sodium channel currents, can solve the problems of low cell harvest rate and survival rate, shortened cell survival time, complicated operation, etc., and achieves good prolongation. state, attenuate spontaneous contractions, and improve the effect of survival rate

Inactive Publication Date: 2012-12-05
辉源生物科技(上海)有限公司
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Problems solved by technology

This traditional separation method has defects such as complicated operation, low cell harvest rate and survival rate; calcium ions are directly canceled in the extracellular perfusate, resulting in significantly shortened cell survival time and other defects, which need to be improved and improved to ensure the accuracy of the experiment. success rate and stability

Method used

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Embodiment Construction

[0028] The method for separating guinea pig ventricular myocytes of the present invention and recording Nav1.5 sodium channel current comprises the following steps:

[0029] 1. Isolation of Single Ventricular Myocytes

[0030] Guinea pigs (220-250 grams in weight) were stunned by a slam on the head, and the heart was quickly removed by opening the chest, and the fat and pericardium were removed in normal Tyrode's solution at 4 degrees Celsius, and the aorta was separated and intubated at 6-8 ml / min Perfuse the Langendorff heart at a rate of 100 mph. First, perfuse with normal Tyrode's solution for 5 minutes to completely flush the residual blood in the heart, then perfuse with calcium-free Tyrode's solution for 5 minutes until the heart stops beating completely, and then use 50 ml of enzymatic hydrolysis solution for 8-10 minutes, until When the heart outflow fluid is obviously drawn, the heart expands and the color becomes lighter, after perfusing with Kraft-Brühe solution f...

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Abstract

The invention provides a method for recording an Nav 1.5 sodium channel current by separating cavy ventricular muscle cells. The method comprises two steps of separating a single cavy ventricular muscle cell and recording the Nav 1.5 sodium channel current. According to the method, the yield rate and the survival rate of the separated cavy ventricular muscle cells are enhanced by improvement of a perfusate and a storage way, and the success rate and the stability of an experiment are increased by the prolonging of the stable time of the cell. The separated cavy ventricular muscle cell has a normal ion channel function and the method is suitable to research on a ventricular muscle physiological characteristic by a patch clamp technique; the experiment operation is easy to master, and the method for acutely separating the ventricular muscle cells is simple and efficient.

Description

technical field [0001] The invention relates to biotechnology, in particular to a method for isolating guinea pig ventricular myocytes and recording Nav1.5 sodium channel current. Background technique [0002] Nav1.5 sodium channel is the main myocardial voltage-gated sodium channel, also known as fast sodium channel. The density of sodium ion channels in human cardiomyocytes is the highest, and the high-density distribution of sodium ion channels is important for the formation and transmission of normal impulses. Changes in the density and dynamics of INav1.5 can interfere with conduction and prolong repolarization. [0003] Studies have shown that LQT syndrome, Brugada syndrome, progressive cardiac conduction defect (progressive cardiac conduction defect, PECD), sick sinus syndrome and other heart diseases are related to cardiac sodium ion channels. [0004] During the development of cardiovascular diseases, observing the antagonism of Nav1.5 sodium channels has become a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N27/26
Inventor 谈学海林佩元陈景才范柳
Owner 辉源生物科技(上海)有限公司
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