Uncovering why a blue fluorescent antibody is so bright
Researchers at Scripps Research Institute in La Jolla, Calif., have teased out why one of a group of fluorescent antibodies fluoresces much more brightly and for a much longer time than others of its kind. The work ultimately could lead to improved sensors for a wide variety of applications, including DNA hybridization assays and mercury sensing.
In 2000, the researchers described the group of fluorescent antibodies, which contained about 10 similar antibodies designed to bind stilbene, a gain medium found in dye lasers. Once complexed with stilbene, the antibodies fluoresce to some degree when excited with UV light between 310 and 355 nm. Their emission spectra range from 362 to 442 nm.
The antibody EP2-19G2-stilbene complex, shown here in crystal form, emits bright blue at 410 nm when excited by UV light. Courtesy of Erik W. Debler.
Within this group, the stilbene complex of antibody EP2-19G2 stands out for its unusually bright fluorescence and long fluorescence lifetime. Its fluorescence is more than an order of magnitude stronger than the other complexes, said Erik W. Debler, lead author of recently published research unraveling why the fluorescence is stronger.
Since its discovery, EP2-19G2 already has had numerous applications. It is used for chiral sensing in high-throughput synthesis systems, to detect mercury, in DNA hybridization assays and to analyze the residues on viral surfaces. Debler said that better characterizing the molecule could expand its uses as a biosensor.
As published in the Feb. 29 issue of Science, the researchers found that the key to the enhanced fluorescence lies in the antibody’s molecular structure. When it binds to stilbene, it organizes its structure to “stack” the stilbene molecule so that it has access to a tryptophan residue at the site of the fluorescence activity. Antibody EP2-19G2 is the only one that has this stacking activity. The result is that this allows stilbene and the tryptophan residue to transfer electrons deep within the molecular complex.
From here Debler said that they plan to continue work on developing additional antibody-chromophore complexes that have charge recombination-induced luminescence phenomena similar to those of EP2-19G2. They also plan to work on further biosensor applications with the existing antibodies.
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