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Method for recording L type calcium channel current by separating guinea pig ventricular muscle cells

A ventricular myocyte and calcium channel technology, applied in biochemical equipment and methods, microbial determination/inspection, etc., can solve the problems of complicated operation, low cell harvest rate and survival rate, shortened cell survival time, etc., to improve the survival rate. rate, prolonging well-being, attenuating the effect of spontaneous contractions

Inactive Publication Date: 2013-05-15
辉源生物科技(上海)有限公司
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AI Technical Summary

Problems solved by technology

This traditional separation method has defects such as complicated operation, low cell harvest rate and survival rate; Na is directly canceled in the extracellular perfusate + , resulting in the obvious shortening of cell survival time and other defects, which need to be improved and improved to ensure the success rate and stability of the experiment

Method used

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Embodiment Construction

[0027] The present invention separates guinea pig ventricular myocyte and records the method for L-type calcium channel electric current, comprises the following steps:

[0028] 1. Isolation of Single Ventricular Myocytes

[0029] Guinea pigs (weight 220-250g) were knocked into a coma by hitting the head, and the heart was quickly taken out by opening the chest. The fat and pericardium were removed in normal Tyrode's solution at 4°C, and the aorta was separated and intubated at 8-10ml / min. Perfuse the isolated heart at a high speed. First, perfuse with normal Tyrode's solution for 5 minutes to completely flush the remaining blood in the heart, then perfuse with calcium-free Tyrode's solution for 5 minutes until the heart stops beating completely, and then use 50ml enzymatic hydrolysis solution for 8-10 minutes, until the heart effluent is obviously drawn, When the heart swells and becomes larger and the color becomes lighter, after perfusion with Kraft-Brühe solution for 5 mi...

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Abstract

An L type calcium channel is mainly distributed in heart, blood vessels, endocrine and nervous systems, and plays a vitally important role in heart and smooth muscle excitation-contraction coupling, atrionector and atrioventricular node action potential conduction. The method that the influence of a compound on the L type calcium channel is observed through a patch clamp recording method so as to evaluate a hearth safety coefficient of the compound and the effect of treating the hypertension becomes an important research mode. According to the method for recording the L type calcium channel current by separating the guinea pig ventricular muscle cells, through improving a perfusate storage mode, the yield rate and the survival rate of separating the guinea pig ventricular muscle cells are increased, and the stabilization time of the cells is prolonged, and thus the success rate and the stability of experiments are increased.

Description

technical field [0001] The invention relates to biotechnology, in particular to a method for isolating guinea pig ventricular myocytes and recording L-type calcium channel current. Background technique [0002] L-type calcium channels are mainly distributed in myocardium, smooth muscle (including blood vessels, intestines, lungs, uterus), endocrine cells (including pancreatic β cells, pituitary) and nerves. L-type calcium channels play an important role in the excitation-contraction coupling of the heart and smooth muscle, the conduction of action potentials in the sinoatrial node and the atrioventricular node, synaptic plasticity, and hormone secretion. [0003] During the development of drugs for hypertension, cardiovascular disease and cancer, observing the antagonism of L-type calcium channels has become an important means to assist in the evaluation of drug effects. The patch clamp technique is known as the "gold standard" for studying ion channels and is the most impo...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/02C12Q1/24
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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