A wide variety of cellular functions depend on the spatial and temporal dynamics of intracellular Ca²⁺ signals. Especially for long lasting processes such as metabolism, gene expression, and cell proliferation, cells rely on the so-called store-operated Ca²⁺ entry pathway, which is particularly prominent in immune cells. Here, sensor proteins (STIM1, STIM2) located within the endoplasmic reticulum (ER) sense the Ca²⁺ concentration in the ER, and upon its depletion, cluster and trap Orai1 proteins into functional regions of the plasma membrane in close apposition to the ER. Highly selective Ca²⁺ channels are opened when the channel-activating domain of STIM proteins bind to the C-terminal of Orai1.

We will discuss the biological background, tools, methodologies and protocols to measure calcium fluxes and downstream effects.  Open questions that benefit from molecular modelling approaches will be discussed.