Parvalbumin, a myoplasmic protein with multiple isoforms in fish muscle, is a low molecular weight protein (9-11 kD) that appears to aid in relaxation from contraction. Parvalbumin aids in relaxation by binding free Ca²⁺, which reduces the intracellular concentration of the ion. Since Ca²⁺ plays a necessary role in contraction by binding to the myofibrillar protein troponin, a decrease in its intracellular concentration will result in relaxation of the muscle fiber. Mechanics measurements have shown that there is a correlation between muscle relaxation and the total amount of parvalbumin as well as the relative expression of the two isoforms (Parv1 and Parv2) in a variety of fish including sheepshead and rainbow trout. Muscle with relatively high relaxation rates expresses more parvalbumin and relatively more of the Parv2 isoform. These physiology results suggest that there are differences in the cation binding properties of each isoform. Since parvalbumin is bound to Mg²⁺ in its native state, Mg²⁺ ions must dissociate for parvalbumin to bind Ca²⁺ ions. Therefore, the dissociation rate for Mg²⁺ is believed to determine the physiological properties of parvalbumin and its ability to aid in relaxation. To test these suggestions we are studying the Ca²⁺ and Mg²⁺ binding characteristics of parvalbumin isoforms in several fish. Here we report on our isolation and characterization of parvalbumin isoforms from sheepshead (Archosargus probatocephalus), rainbow trout (Oncorhynchus mykiss), scup (Stenotomus chrysops), and dogfish shark (Squalus acanthias).