Measurement of Ca²⁺-ATPase activity (in PMCA and SERCA1).

Department of Cardiovascular Sciences, Clinical Division of Anaesthesia Critical, Care and Pain Management, University Hospitals of Leicester NHS Trust, Leicester, Leicestershire, UK. DGL3@le.ac.uk

Methods in molecular biology (Clifton, N.J.) 2013

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Ca(2+)-ATP pumps (those on the plasma membrane; PMCA and sarcoplasmic reticulum; SERCA1) have an important role to play in the regulation of intracellular calcium concentrations. In this chapter, three preparations, two membranes and a purified enzyme, best suited for studies of Ca(2+)-ATPase activity are described. The two selected membranes are the human red blood cell (RBC) ghosts, a representative of plasma membranes (PM), and the rabbit skeletal muscle SR, an intracellular membrane. In this protocol, Pi released during the ATPase reaction is subsequently measured colorimetrically as a complex of molybdovanadate. The method is simple (one-step), fast, sensitive, and reliable.