Technologies

Various detection schemes for spectral properties have been implemented in most commercial confocal microscopes. However, confocal microscopes suffer from at least one of the following problems, when compared to two-photon microscopes: (1) low acquisition speeds, mostly due to use of a point-scan method; (2) photodestruction (bleaching) of fluorescent molecules due to exposure of the whole sample to excitation light (while the signal is only read from a thin layer); (3) low signal-to-noise ratio, due to the fact that excitation and emission occur at wavelengths close to one another, making filtering of the signal difficult. On the other hand, two-photon microscopes, which avoid problems (2) and (3) above, either do not present spectral resolution at all, or, when they do, they suffer from slow acquisition speed. Dr. Raicu’s technology produces spectrally resolved fluorescence images of samples using a two-photon microscope after only one full scan of the sample, which avoids all of the aforementioned problems.

Laser scanning microscopes (such as two-photon and confocal microscopes) are widely used for acquiring images of narrow sections of cells and tissues, in which molecules of interest are tagged with fluorescent molecules. Fluorescent tags make the molecules of interest visible through light emission. By acquiring multiple such thin sections, three-dimensional images of the samples can be obtained. In experiments involving fluorescence tagging with multiple colors, such as in studies of protein colocalization or protein-protein interactions, a fourth dimension becomes critically necessary: the spectral dimension (i.e., the wavelength of the emitted light).