![]() The type of system employed, whether it be cell culture or whole animal, the depth of imaging, the relevant time period available for data collection, and even the ability to distinguish multiple reporter systems from within the same tissue must be understood and acknowledged prior to beginning any experiment. As a result, much consideration must be given to the experimental design associated with bioluminescent or fluorescent detection from mammalian cells. The differences in gene expression and cellular compartmentalization between prokaryotic and eukaryotic cells represent the major obstacles for the efficient expression of these and other reporter systems at the cellular level, but once the line has been crossed from expression in single cells to expression in multicelluar organisms, these problems can be compounded by the increases in absorption and scattering intrinsic to whole animal imaging. ![]() However, the expression and detection of these reporter systems in eukaryotic models presents unique challenges not encountered in their prokaryotic counterparts. This has occurred in part because the use of cultured mammalian cells or small animal models has increased steadily over time in order to obtain more relevant human proxies for the measurement of cellular processes and bioavailability of biomedically relevant compounds of interest. While originally utilized primarily in prokaryotic organisms, reporter systems such as green fluorescent protein (GFP) and its variants, substrate dependent luciferase systems such as beetle and marine luciferase proteins, and substrate independent luciferase systems such as the bacterial luciferase gene cassette have now become the standards for imaging in the mammalian cellular background as well ( Fig.
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