Nature Methods 3, 259 - 261 (2006)
Published online: 22 March 2006; | doi:10.1038/nmeth862
Imaging specific cell-surface proteolytic activity in single living cellsJohn P Hobson1, Shihui Liu2, Birgitte Rønø1, 3, Stephen H Leppla2
& Thomas H Bugge11
Proteases and Tissue Remodeling Unit, National Institute of Dental and Craniofacial Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 30 Convent Drive, Bethesda, Maryland 20892, USA. 2
Microbial Pathogenesis Section, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 30 Convent Drive, Bethesda, Maryland 20892, USA. 3
Finsen Laboratory, Strandboulevarden 49, DK-2100 Copenhagen Ø, Denmark.
Correspondence should be addressed to Stephen H Leppla leppla@nih.gov or Thomas H Bugge thomas.bugge@nih.gov We describe a simple, sensitive and noninvasive assay that uses nontoxic, reengineered anthrax toxin– -lactamase fusion proteins with altered protease cleavage specificity to visualize specific cell-surface proteolytic activity in single living cells. The assay could be used to specifically image endogenous cell-surface furin, urokinase plasminogen activator and metalloprotease activity. We have adapted the assay for fluorescence microscopy, flow cytometry and fluorescent plate reader formats, and it is amenable for automation and high-throughput analysis.
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