Current and prospective trends in the application of bioluminescent analysis in experimental brain studies Elena N. Esimbekova a,b , Valentina A. Kratasyuk a,b , Natalia A. Rozanova c,d , Victoria I. Lonshakova-Mukina a,b , Irina G. Torgashina a , Yulia K. Komleva c,d , Mikis R. Saridis d , Sofia A. Korsakova d , Stanislav O. Yurchenko d , Alla B. Salmina c,d,* a Siberian Federal University, Svobodny ave., 79, Krasnoyarsk, Russia b Institute of Biophysics, Siberian Branch of Russian Academy of Sciences, Akademgorodok, Krasnoyarsk, Russia c Brain Science Institute, Russian Center of Neurology and Neurosciences, Volokolamskoe Highway, 80, Moscow, Russia d Centre for Soft Matter and Physics of Fluids, Bauman Moscow State Technical University, 2nd Baumanskaya str., 5, Moscow, Russia A R T I C L E INFO Keywords: Bioluminescence Bioluminescent assay Luciferase D-luciferin Coelenterazine Brain imaging Neurobiology ABSTRACT Bioluminescence, a natural phenomenon resulting from enzyme-catalysed oxidation of substrates like luciferin, has emerged as a powerful tool in modern biology, biochemistry, and medicine. Advances in understanding bioluminescent systems, particularly those of bacteria, fireflies, coelenterates, and crustaceans, have facilitated the development of highly sensitive and specific analytical methods. Applications range from intracellular bio- imaging to the detection of metabolites and effectors, offering unparalleled insight into cellular and molecular processes. Recently, experimental neuroscience has embraced bioluminescent techniques, leveraging their unique advantages for visualizing and assessing brain cell function and metabolic activity in vivo and in vitro. Furthermore, the potential to control neuronal activity through bioluminescent reactions is opening new fron- tiers in neurobiology. This review focuses on the principles of bioluminescent assays based on bacterial and firefly luciferases and coelenterazine-derived systems, highlighting their current and prospective applications in experimental brain studies. 1. Introduction Wide comprehensive studies of bioluminescence phenomena have allowed the development of many practical bioluminescence techniques for application in various fields of biology, biochemistry, biotechnology, and medicine. Currently, the most studied bioluminescent systems are those of bacteria, fireflies, coelenterates and crustaceans. Despite the single end result of bioluminescence – the emission of light as a product of the reaction of oxygen oxidation of the substrate, luciferin, catalysed by a specific enzyme, luciferase – the biochemical mechanisms of bioluminescence are quite different. The names luciferin and luciferase, originally used to describe the bioluminescent reaction of fireflies, are now collectives, and the bioluminescent enzymes and substrates of different organisms have been given their specific names. When the generic terms ‘luciferin’ and ‘luciferase’ are used, it is customary to be sure to stipulate their specificity, as they refer to completely different chemical compounds and proteins. Considerable research effort was devoted to bioluminescence mechanisms in the 1960–90s, providing the basis for the creation of a variety of analytical methods. First of all, of course, it is necessary to note the enormous success of using bioluminescence systems in bio- imaging of intracellular processes in vivo. Recombinant proteins ob- tained by cloning genes of bioluminescent proteins are used to solve many problems where visualization of ongoing processes is required – from studying protein-protein interactions in individual cells to behav- ioural reactions of organisms. Another, no less wide sphere of applica- tion of bioluminescent systems is analytical tasks of identification of separate compounds (metabolites, inhibitors, effectors), which is possible due to high sensitivity and specificity of bioluminescent methods. An interesting and relatively new application of bioluminescence analysis is experimental neuroscience. Visualization of brain cells and assessment of their functional or metabolic activity using biolumines- cence is becoming increasingly demanded in in vivo and in vitro studies. * Corresponding author. Brain Science Institute, Research Center of Neurology, Volokolamskoe Highway, 80, Moscow, Russia. E-mail address: allasalmina@mail.ru (A.B. Salmina). Contents lists available at ScienceDirect Talanta journal homepage: www.elsevier.com/locate/talanta https://doi.org/10.1016/j.talanta.2025.128510 Received 15 April 2025; Received in revised form 6 June 2025; Accepted 22 June 2025 Talanta 296 (2026) 128510 Available online 24 June 2025 0039-9140/© 2025 Elsevier B.V. All rights are reserved, including those for text and data mining, AI training, and similar technologies.