In vivo imaging of beta-galactosidase activity using far red fluorescent switch.
beta-Galactosidase (beta-gal) has been widely used as a transgene reporter enzyme, and several substrates are available for its in vitro detection. The ability to image beta-gal expression in living animals would further extend the use of this reporter. Here we show that DDAOG, a conjugate of beta-galactoside and 7-hydroxy-9H-(1,3-dichloro-9,9-dimethylacridin-2-one) (DDAO), is not only a chromogenic beta-gal substrate but that the cleavage product has far-red fluorescence properties detectable by imaging. Importantly, the cleavage substrate shows a 50-nm red shift, enabling its specific detection in a background of intact probe, a highly desirable feature for in vivo imaging. Specifically, we show that beta-gal-expressing 9L gliomas are readily detectable by red fluorescence imaging in comparison with the native 9L gliomas. We furthermore show that herpes simplex virus amplicon-mediated LacZ gene transfer into tumors can be transiently and thus serially visualized over time. The results indicated that in vivo real-time detection of beta-gal activity is possible by fluorescence imaging technology.
Center for Molecular Imaging Research, Massachusetts General Hospital, Harvard Medical School, Charlestown, MA 02129, USA. tung@helix.mgh.harvard.edu
Ching-Hsuan
CH
Tung
Qing
Q
Zeng
Khalid
K
Shah
Dong-Eog
DE
Kim
Dawid
D
Schellingerhout
Ralph
R
Weissleder
D000818:Animals; D005456:Fluorescent Dyes; D022761:Gene Transfer, Horizontal; D018259:Herpesvirus 1, Human / Q000235:genetics; D007763:Lac Operon; D051379:Mice; D012680:Sensitivity and Specificity; D001616:beta-Galactosidase / Q000032:analysis*
D005456:Fluorescent Dyes; D001616:beta-Galactosidase