Physiologically, when actin and myosin combine the muscle produces force. There are several methods to measure the stoichiometry of actin to myosin combination. M4 shows, without actin only 1 peak, the myosin peak, is seen in the analytical ultracentrifuge. When all the myosin combined with actin, again only 1 peak, the actomyosin peak, is seen. Myosin-actin binding as followed in the analytical ultracentrifuge.
Viscometry is another method to measure myosin-actin binding in solution.
The viscosity of actomyosin is much higher than the sum of its individual components actin and myosin. Thus, when constant amount of myosin is titrated with increasing amount of fibrous actin the stoichiometry of the actin binding is reached at the maximal viscosity. In suspension, at physiological ionic strength, the myosin-actin binding is measured under conditions when both myosin and actomyosin are insoluble, that is they sediment at low centrifugal fields. Thus, adding increasing amount of fibrous actin to constant amount of myosin in suspension will result in actomyosin formation. After myosin becomes saturated with actin, the uncombined actin remains in the supernatant of the centrifugate.