Hello,

I have tried the attached protocol to isolate primary hepatocyte from mice, using Collagenase instead of Liberase.

Everything is the same as mentioned in the protocol. However, by the time the liver is dissected the cells are all dead.

• The pH and temperature of the buffer is 7.4 and 37C respectively.
• Cannulation goes well and liver gets digested
• Used isoflurane anesthesia 5% for induction and 1.3% for maintenance
• perfusion flow rate :7ml/min
• The cells are all dead immediately after harvesting liver

I want to open it up to a discussion because I am desperately finding out what exactly is going wrong for such low viability.