I have subjected pieces of explanted liver samples for collagenase treatment (1/2 an hour at 37), and purified through Percoll centrifugation. However, the cells I obtained do not look polygonal, but rather are much rounder and very small (lymphocyte like). I would like to obtain a protocol for isolation of hepatocytes that works robustly with liver pieces (many protocols I see call for in situ collagenase perfusion in rats/mice, which is not possible for human samples).
Hi
A paper published in JoVE show the protocol about isolation of human hepatocytes by a two-step collagenase perfusion procedure. They also provide video and detail information. Please check this website http://www.jove.com/video/50615/isolation-human-hepatocytes-two-step-collagenase-perfusion
Hi
A paper published in JoVE show the protocol about isolation of human hepatocytes by a two-step collagenase perfusion procedure. They also provide video and detail information. Please check this website http://www.jove.com/video/50615/isolation-human-hepatocytes-two-step-collagenase-perfusion
I recently was having this problem with hepatocytes differentiated from hESCs. It might be that your cells are clumping together, i found that DNA from disrupted cells can cause gooey strings to form which prevents cells filtering efficiently. Try adding DNAse to your protocol, I found Benzonase to be very efficient. I can send you the whole protocol if you like.
- Badges
- Science topic
- Similar topics
- Methodology
- Laboratory
- Cellular Biology Techniques