Bioluminescent imaging is an emerging biomedical surveillance strategy that uses external cameras to detect in vivo light generated in small animal models of human physiology or in vitro light generated in tissue culture or tissue scaffold mimics of human anatomy. The most widely utilized of reporters is the firefly luciferase (luc) gene; however, it generates light only upon addition of a chemical substrate, thus only generating intermittent single time point data snapshots. To overcome this disadvantage, we have demonstrated substrate-independent bioluminescent imaging using an optimized bacterial bioluminescence (lux) system. The lux reporter produces bioluminescence autonomously using components found naturally within the cell, thereby allowing imaging to occur continuously and in real-time over the lifetime of the host. We have validated this technology in human cells with demonstrated chemical toxicological profiling against exotoxin exposures at signal strengths comparable to existing luc systems (~1.33 × 107 photons/second). As a proof-in-principle demonstration, we have engineered breast carcinoma cells to express bioluminescence for real-time screening of endocrine disrupting chemicals and validated detection of 17β-estradiol (EC50 = ~ 10 pM). These and other applications of this new reporter technology will be discussed as potential new pathways towards improved models of target chemical bioavailability, toxicology, efficacy, and human safety.
Bioluminescent production represents a facile method for bioreporter detection in mammalian tissues. The lack of
endogenous bioluminescent reactions in these tissues allows for high signal to noise ratios even at low signal strength
compared to fluorescent signal detection. While the luciferase enzymes commonly employed for bioluminescent
detection are those from class Insecta (firefly and click beetle luciferases), these are handicapped in that they require
concurrent administration of a luciferin compound to elicit a bioluminescent signal. The bacterial luciferase (lux) gene
cassette offers the advantages common to other bioluminescent proteins, but is simultaneously capable of synthesizing
its own luciferin substrates using endogenously available cellular compounds. The longstanding shortcoming of the lux
cassette has been its recalcitrance to function in the mammalian cellular environment. This paper will present an
overview of the work completed to date to overcome this limitation and provide examples of mammalian lux-based
bioreporter technologies that could provide the framework for advanced, biomedically relevant real-time sensor
development.
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