Myocardial cell separation method

A separation method and cardiomyocyte technology, applied in cell dissociation methods, animal cells, vertebrate cells, etc., can solve the problems of reducing the survival rate of cardiomyocytes, deformation of cardiomyocytes, etc., and achieve the effect of avoiding changes and improving activity

Inactive Publication Date: 2017-05-17
HENAN MEDICAL COLLEGE
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0004] The method for separating cardiomyocytes provided by the present invention solves the problems in the prior art t

Method used

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  • Myocardial cell separation method

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Embodiment 1

[0041] A method for separating cardiomyocytes of adult rats, comprising the following steps:

[0042] Step 1: Put the freshly isolated heart into the pre-cooled protective solution at 4°C, cut out the ventricular muscle tissue, wash it with washing solution, and then cut the ventricular muscle tissue into small pieces of 1mm×1mm×1mm with ophthalmic scissors , then soaked in the flushing solution for 3 minutes, filtered with gauze to obtain a myocardial tissue sample, and set aside;

[0043] Each liter of the protective solution contains the following components: 8g of NaCl, 0.4g of KCl, 0.06g of Na 2 HPO 4 ·H2 O, 0.06 g of KH 2 PO 4 , 0.35g of NaHCO 3 , the solvent is double distilled water;

[0044] Each liter of washing solution contains the following components: 9g of NaCl, 0.3g of KCl, and the solvent is double distilled water.

[0045] Step 2, the myocardial tissue sample was placed in 1 g / L trypsin solution, digested at 37° C. for 8 min, and the supernatant was dis...

Embodiment 2

[0051] A method for separating cardiomyocytes from neonatal rats, comprising the following steps:

[0052] Step 1: Put the freshly isolated heart into the pre-cooled protective solution at 4°C, cut out the ventricular muscle tissue, wash it with washing solution, and then cut the ventricular muscle tissue into small pieces of 1mm×1mm×1mm with ophthalmic scissors , then soaked in the flushing solution for 5 minutes, filtered with gauze to obtain a myocardial tissue sample, and set aside;

[0053] Each liter of the protective solution contains the following components: 8g of NaCl, 0.4g of KCl, 0.06g of Na 2 HPO 4 ·H 2 O, 0.06 g of KH 2 PO 4 , 0.35g of NaHCO 3 , the solvent is double distilled water;

[0054] Each liter of washing solution contains the following components: 9g of NaCl, 0.3g of KCl, and the solvent is double distilled water.

[0055] Step 2, the myocardial tissue sample was placed in 1 g / L trypsin solution, digested at 37° C. for 5 min, and the supernatant ...

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Abstract

The invention discloses a myocardial cell separation method, comprising the following steps: putting a fresh excised heart into 4DEG C precooled protection fluid, scissoring ventricular tissues, washing the ventricular tissues with flushing fluid, and then scissoring the ventricular tissues into bits; then sequentially carrying out trypsin digestion, high concentration buffer immersion, digestive juice digestion and low concentration buffer immersion; uniformly mixing a mixture of the high concentration buffer immersion fluid, postdigestive supernatant fluid and low concentration buffer immersion fluid, beating with a suction tube, then filtering off myocardial tissue precipitate, collecting the filtrate and centrifuging to obtain the separated myocardial cell precipitate. The method disclosed by the invention uses concentration decrease of potassium ions to generate stimulation on the myocardial cells, thereby improving the activity of the myocardial cells, and avoiding the problems of change of form of the myocardial cells and reduction of survival rate of the cells.

Description

technical field [0001] The invention belongs to the technical field of cardiomyocyte separation, and in particular relates to a cardiomyocyte separation method. Background technique [0002] Cardiomyocytes are one of the important cells in the growth, development, physiology and metabolism of animals. In order to facilitate the study of the physiological function of cardiomyocytes, observe the cell morphology, and eliminate the influence of various limiting factors on cardiomyocytes in vivo, in vitro isolation and culture methods are usually used. [0003] Since cardiomyocytes are located in the myocardial tissue of the heart, experimenters usually need to inject digestive fluid into the isolated heart when isolating cardiomyocytes, and collect the cardiomyocytes after the digestion is complete. However, when the cardiomyocytes are isolated, the heart has already been separated from the body and lacks protection. Prolonged digestion can easily cause deformation of the cardi...

Claims

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Application Information

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IPC IPC(8): C12N5/077
CPCC12N5/0657C12N2509/00
Inventor 刘芳韩圣娜李宁宁陈文超师瑞红
Owner HENAN MEDICAL COLLEGE
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