Issue Date: April 7, 2014 | Web Date: April 3, 2014
Better Biomolecular Visualization
Bioorthogonal reactions—reactions in cells that don’t interfere with life processes—have made it possible in recent years to measure and visualize specific biomolecules in living cells and organisms. But the technology is sometimes difficult to carry out, especially in live animals.
Grad student Peyton Shieh, chemistry professor Carolyn Bertozzi, and coworkers at the University of California, Berkeley, have now developed a new bioorthogonal reaction that overcomes key problems (Proc. Natl. Acad. Sci. USA 2014, DOI: 10.1073/pnas.1322727111).
They used computation to develop a new fluorogenic reagent—a reactant whose fluorescence is significantly enhanced after it reacts, in this case with a labeled biomolecule inside a living cell. The boosted fluorescence is then detected from outside the cell to track the biomolecule.
Bioorthogonal reactions generally produce visible-wavelength fluorescence, but the new reaction produces near-infrared radiation. Near-IR wavelengths penetrate tissues more readily than visible radiation and are less prone to interferences.
In the approach, a fluorogenic azide reacts with a cyclooctyne group on a metabolically labeled biomolecule in a fast, highly efficient click chemistry reaction. The reaction doesn’t require a catalyst, making it more benign than click chemistry reactions catalyzed by copper, which is toxic to cells.
The product’s fluorescence is enhanced (relative to that of the reagent) about 50 times, compared with less than a factor of two enhancement for previous bioorthogonal fluorogenic reagents. This obviates the need to remove excess unreacted reagent from cells as is typically required, especially when nonfluorogenic reagents are used. “The new capability these probes bring to the table is imaging in live animals, where washing out unreacted probe is essentially impossible,” Bertozzi says. Her group used the approach to visualize glycoproteins on mammalian cells and peptidoglycans in live bacteria.
“Minimizing nonspecific background and generating good contrast is the secret of success, so this is a good approach,” comments imaging specialist Kevin M. Brindle of Cambridge University.
- Chemical & Engineering News
- ISSN 0009-2347
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