SUMMARYIn this review, we take a retrospective look at the discovery and utilization of the Ca2+‐sensitive bioluminescent protein complex, aequorin. We do consider the contribution it has made to our understanding of the natural phenomenon of bioluminescence, but it is in the application of extracted and purified aequorin as a reporter of Ca2+dynamics in living cells, which is arguably its major contribution to biological and biomedical science. Following its extraction, purification, and subsequent availability in the mid‐1960s, aequorin became the intracellular reporter of choice until it was replaced in the late 1970s by easier‐to‐use fluorescence‐based reporters. From the mid‐1980s onwards, however, aequorin‐based Ca2+imaging underwent a renaissance following the cloning of the aequorin gene and the emergence of routine techniques to target and express it exogenously in plant and animal systems. The development of aequorin as a tool continues as spectral varieties are being developed that allow simultaneous imaging of Ca2+dynamics in different cellular organelles and microdomains. We predict that further developments in the use of aequorin, as well as other bioluminescent proteins, will continue, especially in the areas of regenerative medicine and whole organism imaging.Mol. Reprod. Dev. 82: 563–586, 2015. © 2014 Wiley Periodicals, Inc.