Calcium uptake in crude tissue preparation

The various isoforms of the sarco/endoplasmic reticulum Ca²⁺ ATPase (SERCA) are responsible for the Ca²⁺ uptake from the cytosol into the endoplasmic or sarcoplasmic reticulum (ER/SR). In some tissues, the activity of SERCA can be modulated by binding partners, such as phospholamban and sarcolipin. The activity of SERCA can be characterized by its apparent affinity for Ca²⁺ as well as maximal enzymatic velocity. Both parameters can be effectively determined by the protocol described here. Specifically, we describe the measurement of the rate of oxalate-facilitated ⁴⁵Ca uptake into the SR of crude mouse ventricular homogenates. This protocol can easily be adapted for different tissues and animal models as well as cultured cells.