Hi,
I am trying to harvest hepatocytes from high fat fed C57BL/6 mice. Because of some genetic manipulation and high fat diet, they are actually pretty obese. I find it difficult to make the IV channel to vena cava as its almost covered by visceral fats. Also it seems like even after proper channel the livers are not digesting well. As a result I am getting clumpy liver tissues and the hepatocyte yield is very low. Should I increase the collagenase concentration in the digestion buffer? But my fear is that might kill the cells. The livers are clearly having hepatic steatosis in this group. I have no issues in making nice IV channel or getting hepatocytes from regular chow fed group. Any insights will be highly appreciative.
Hello,
Please see if this paper (from a past colleague of mine) is helpful:
Article Hepatic Sel1L‐Hrd1 ER‐associated degradation (ERAD) manages ...
Read the Primary Hepatocyte Culture under Materials and Methods.
Best,
Anoop
Anoop Arunagiri This is very helpful. What I realize I was using almost half of the collagenase amount, so may be that was not enough for the digestion especially mice with fatty liver and fibrosis. Thanks. I would increase the collagenase amount and will see the cell count.
Thanks.
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