The aequorin complex comprises a 22,000-dalton apoaequorin protein, molecular oxygen and the luminophore coelenterazine. When three Ca2+ ions bind to this complex, coelenterazine is oxidized to coelenteramide, with a concomitant release of carbon dioxide and blue light. The approximately third-power dependence of aequorin's bioluminescence on Ca2+ concentration allows the measurement of Ca2+ concentrations with a broad detection range from ~0.1 µM to >100 µM. Unlike fluorescent Ca2+ indicators, Ca2+-bound aequorin can be detected without illuminating the sample, thereby eliminating interference from autofluorescence.
AnaSpec offers coelenterazine and several synthetic coelenterazine analogs for reconstituting aequorin in cells that have been transfected with apoaequorin cDNA. In addition to native coelenterazine, we also offer a few derivatives of coelenterazine that confer different Ca2+ affinities and spectral properties on the aequorin complex. Recombinant apoaequorin reconstituted with coelenterazine hcp is reported to have the best luminescence overall, with both a high quantum yield and a fast response time. However, intracellular reconstitution of aequorin from coelenterazine analogs can be relatively slow. Aequorins containing the cp, f or h form of coelenterazine exhibit 10–20 times stronger luminescence than that of apoaequorin reconstituted with native coelenterazine. Coelenterazine cp has been used in HTS screening assay for GPCRs.