Table 1.

Frequent causes unsuccessful hepatocyte isolation.

Step	Problem	Possible Reason	Solution
Perfusion	Poor cell dissociation	Poor digestion	Make sure the Digestion solution is at 37°C
		Tissue disruption	Lower down perfusion speed
	Cell death	Excessive collagenase digestion	Reduce the time/volume of perfusion with Hepatocyte Liver Digest Medium
Cell isolation	Low recovery	To many cells loaded on the gradient	If using more than three lobes, increase the number of gradient tubes
		Gradient disturbance	Setup new gradient, making sure the layers do not mix
		Altered gradient densities	Check that the gradient centrifugation temperature is kept at 20°C and Percoll solutions at room temperature