Method for separating primary adult hepatocytes, and special sterile apparatus box thereof
A hepatocyte and instrument box technology, applied in artificial cell constructs, animal cells, vertebrate cells, etc., can solve the problems of unable to meet the huge demand of human hepatocytes, unable to implement liver perfusion, and difficult to find perfusion blood vessels, etc. The effect of reducing tedious preparation work, simple structure and convenient use
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Embodiment 1
[0037] The setting of embodiment one disposable aseptic instrument box
[0038] figure 1 An embodiment of the disposable kit for isolating primary human hepatocytes shown is composed of an outer packaging bag 1 , a box body 4 and operating instruments, and the box body 4 is set in the outer packaging bag 1 . Operating equipment includes 10cm sterile Petri dish 2, 100ml sterile beaker 7, 20ml sterile beaker 8, 25ml sterile serum bottle 3, 50ml sterile triangular beaker 9 and 50ml sterile triangular beaker 10, 200 mesh sterile nylon filter 14 and 100 mesh sterile nylon filters 5, 10ml sterile syringes 15 and 20ml sterile syringes 11, sterile forceps 6 and sterile scissors 13, two 15ml centrifuge tubes 12. The box body 4 is provided with a plurality of grooves matching the shape and volume of the above-mentioned operating instruments, and each of the above-mentioned operating instruments is correspondingly embedded in the grooves in the corresponding box body 4 .
[0039] These...
Embodiment 2
[0040] Example 2 Isolation and Culture of Primary Adult Hepatocytes
[0041] DMEM medium: prepared according to the formula in "Cell Culture" (Situ Zhenqiang, Wu Junzheng editor-in-chief. Xi'an: World Book Publishing Company, 2007. pp. 50-52).
[0042] The composition of preperfusate contains NaCl 8g / L, KCl 0.4g / L, NaCl 2 HPO 4 12H 2 O 0.12g / L, KH 2 PO 4 0.06g / L, NaHCO 3 0.35g / L, glucose 1.0g / L, HEPES 2.38g / L, EDTA 0.74g / L.
[0043] The type IV collagenase solution is prepared by dissolving the type IV collagenase in DMEM medium, and the concentration of the type IV collagenase is 0.05-0.1%. The amount of type IV collagenase solution can be determined according to the size and weight of liver tissue. However, in order to save resources, it has been proved by experiments that 15ml of type IV collagenase solution is used when the weight of liver tissue is 1.0-3.0g, and 20ml of IV collagenase solution is used when the weight of liver tissue is 3.1g-4.0g. Type IV collagena...
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