Hi all,

I am currently working on rat hepatocyte isolation.I use HBSS(Ca2+ free) from sigma(9.5g/L) as perfusion medium via cannulation of portal vein.Then in about 2 min once the liver gets cleared of RBCs as evidenced by the change in color.I pass collagenase solution(0.05% collagenase type IV sigma),this solution also consists of 0.018%CaCL2, for digestion for about 6 min.After excising the liver I filter it through 100 um cell strainer,then centrifuge at 430 RPM.But the problem is sometimes I get a coagulated mass of cells.This happens rarely but I couldn't figure out the reason. So,if anyone has any experience in this area any help or suggestions would be appreciated.Thanks.