Problem	Solution
Low quality of mitochondria	The animals are normally deprived of food overnight to reduce the glycogen content of the liver; this facilitates the separation process.
Low respiratory control	The clearance between the glass wall and the side of a pestle must be 0.3 – 0.5 mm. Use of a pestle that is too tight, or excessive homogenization, will damage the mitochondria. High yields and quality also depend on the gentle and rapid handling of the tissue. For a whole liver, divide the starting material into 2–3 portions and homogenize them separately. Removal of this lipid is essential, as free fatty acids are potent uncouplers.
Low purity of isolated mitochondria	Contamination with other organelles can be reduced by washing the mitochondrial pellet gently.