Separation and culture method of renal podocyte of mouse

Abstract

The invention provides a separation and culture method of renal podocyte of a mouse. The separation and culture method comprises the following steps of (a), intraperitoneally injecting 2% pentobarbital sodium to execute the mouse, sterilizing the mouse with alcohol, taking out kidney tissue of the mouse, putting the kidney tissue into a precooled sterile PBS (Phosphate Buffer solution) containing double antibodies, soaking and washing the kidney tissue, and peeling and removing a renal capsule; (b), mechanically dissociating the kidney tissue, then digesting the kidney tissue by a preheated mixed enzyme solution, and terminating digestion by a mixed complete culture solution; (c), obtaining the renal podocyte by adopting a difference sieving method, inoculating the podocyte in a treated cell bottle, and culturing the podocyte; (d), digesting a cell, which is cultured for 3d, through pancreatin, then filtering the cell once again by a sieve mesh, centrifuging filtrate to remove supernatant fluid, and resuspending and inoculating an obtained mixture in the treated cell culture flask; (e), identifying the renal podocyte by cell immunofluorescence. The separation and culture method of the renal podocyte of the mouse, which is provided by the invention, is good in repeatability, and is simple and convenient to operate; moreover, the high-yield and high-purity renal podocyte of the mouse can be obtained.

Description

technical field [0001] The invention belongs to the field of cell biology, in particular to a method for separating and culturing mouse renal podocytes. Background technique [0002] Podocytes, the visceral epithelial cells of the renal capsule, attach to the outermost side of the glomerular basement membrane (GBM) and together with the glomerular endothelial cells and capillary endothelium constitute the renal microvessels. The molecular barrier and charge barrier of the spherical filtration membrane, the slit diaphragm formed between the foot processes is the last barrier of the filtration membrane. Podocyte damage plays an important role in the development of proteinuria, and many glomerular diseases accompanied by podocyte damage may progress to chronic renal failure. The research on the physiological characteristics of podocytes and their injury response mechanism can further understand the pathogenesis of proteinuria and glomerular diseases. However, because renal po...

AI Technical Summary

Problems solved by technology

However, because renal podocytes are terminally differentiated cells with complex structures and special positioning, primary cells cultured in vitro can hardly proliferate, and the cell genome of immortalized podocyte lines changes, which cannot accurately reflect renal physiology and pathology Therefore, the present invention aims to provide a method for isolating and culturing mouse renal podocytes with good repeatability and easy operation, and can obtain mouse renal podocytes with high yield and high purity. Provide a reasonable experimental basis for studying the mechanism of podocyte injury

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