Efficient separation and purification method of sheep skeletal muscle satellite cells

Abstract

The invention discloses an efficient separation and purification method of sheep skeletal muscle satellite cells, which comprises the following steps: soaking muscle tissues in 75% alcohol for 15 minutes, cutting required skeletal muscles, cleaning with PBS (Phosphate Buffer Solution) containing 1% double antibodies, removing connective tissues, cutting the skeletal muscles into 2 mm<3>, transferring the skeletal muscles into a centrifugal tube, repeatedly blowing and beating with the PBS for three times, carrying out natural sedimentation, discarding supernate, transferring tissue blocks into a culture bottle, absorbing the redundant liquid, adding 1ml of culture medium per 25cm<2> growth surface, slowly inclining the culture bottle until tissue blocks are uniformly distributed on the growth surface, covering a bottle cap, culturing at 37 DEG C, 5% CO2 and saturated humidity for 8-24 hours, then adding 1ml of culture medium, continuously adding 1ml of culture medium every 24 hours until the volume of the culture medium is 5ml / 25cm<2>, and changing the culture medium once a week until the cell growth tends to be stable. The method disclosed by the invention is low in cost and extremely small in cell damage, so that more cells can be collected.

Description

Technical field [0001] The present invention relates to the field of biotechnology, and more particularly to an efficient separation and purification method of sheep skeletal muscle satellite cells. Background technique [0002] Skeletal muscle satellite cells (SMSCs) are adult stem cells, which are also coated with skeletal muscle growth and repairing theorematic precursor cells, which are located between the basement film and the base film of the muscle fibers. Due to its unique presence, the separation process of skeletal muscle satellite cells is different from the separation and purification of other cells such as muscle fibroblasts, compared to other cells, the separation method of skeletal muscle satellite cells is more complex and more difficult. Furthermore, other cells are typically doped during the SMSCS separation, such as fibroblasts, endothelial cells, and other blood vessels, etc., so if the pure skeletal muscle satellite cells are obtained, the scientific and reas...

AI Technical Summary

Problems solved by technology

[0005] The enzymes (such as collagenase, trypsin, etc.) used in the commonly used two-step enzymatic digestion and separation of skeletal muscle satellite cells have a greater damage to the cells, so the number of cells obtained by the two-step enzymatic method is often small or growth retardation

The commonly used cell purification methods include differential attachment method and percoll gradient centrifugation, etc. The centrifugation process can cause serious damage to cells, thereby reducing the efficiency of cell collection.

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