Non-myofibrillar enzyme activities persist after glycerination of insect flight muscle. Preservation of mitochondrial structure and function in glycerol-extracted preparations complicates the study of direct effects on the myofibrillar contractile machinery. Non-myofibrillar adenosine triphosphatase and a significant amount of respiration and oxidative phosphorylation in the presence of added substrate persist in glycerol-extracted fibres of insect fibrillar muscle stored for 6 months at -18°C. The standard procedure used to extract actomyosin from myofibrils also extracts from the mitochondrial fraction a protein which shows the phenomenon of superprecipitation and undergoes viscosity changes in the presence of adenosine triphosphate and metal ions. Preparation of the true contractile structure of insect flight muscle therefore requires the removal of all non-myofibrillar activities. Reduction in the total number of mitochondria and of non-myofibrillar enzymic activity is achieved by treatment with the detergent Tween 80 subsequent to glycerination without impairing the mechanical performance of the fibres. This provides better experimental material for studies of the contractile system than the preparations commonly used. The fact that fibres can still perform oscillatory work after treatment with detergent and oligomycin confirms that the control mechanism for oscillation resides within the myofibrils.

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