Optimization of preparation of mitochondria from 25-100 mg skeletal muscle.

A method for isolation of mitochondria from 25-100 mg skeletal muscle is described. The instrumental developments include a refined homogenization setup, special handling techniques, and equipment for biopsy storage. The preparation medium was a standard ionic medium. All fractions were assayed for marker enzymes and the data used in optimization of the yield. It was observed that the homogenization procedure exerts strong control on the integrity of the isolated mitochondria. The method was developed with pigeon breast muscle as the model tissue and used virtually unaltered for preparation from muscles of pigs, rats, and humans. The relative yield was 40-50% and the mitochondria were well coupled and showed high rates of phosphorylating respiration.