Nature Methods 3, 295 - 301 (2006)
Published online: 22 March 2006; | doi:10.1038/nmeth868
Luminescent imaging of  -galactosidase activity in living subjects using sequential reporter-enzyme luminescenceThomas S Wehrman1, 3, Georges von Degenfeld1, 3, Peter O Krutzik1, 2, Garry P Nolan1
& Helen M Blau1, 21
Baxter Laboratory in Genetic Pharmacology, Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, California 94305, USA. 2
Department of Molecular Pharmacology, Stanford University School of Medicine, Stanford, California 94305, USA. 3
These authors contributed equally to this work.
Correspondence should be addressed to Helen M Blau hblau@stanford.edu We generated a sequential reporter-enzyme luminescence (SRL) technology for in vivo detection of  -galactosidase (-gal) activity. The substrate, a caged D-luciferin–galactoside conjugate, must first be cleaved by -gal before it can be catalyzed by firefly luciferase (FLuc) to generate light. As a result, luminescence is dependent on -gal activity. Using this technology, constitutive -gal activity in engineered cells and inducible tissue-specific -gal expression in transgenic mice can now be visualized noninvasively over time. A substantial advantage of -gal as a bioluminescent probe is that the enzyme retains full activity outside of cells, unlike FLuc, which requires intracellular cofactors. As a result, antibodies conjugated to the recombinant -gal enzyme can be used to detect endogenous cells and extracellular antigens in vivo. Thus, coupling the properties of FLuc to the advantages of -gal permits bioluminescent imaging applications that previously were not possible.
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