The most commonly used fluorophores have excitation spectra in the 400–500 nm
range, whereas the laser used to excite the fluorophores lies in the ~700–1000 nm (infrared) range. If the fluorophore absorbs two infrared photons simultaneously, it will absorb enough energy to be raised into the excited state.
Therefore,
The fluorophore will then emit a single photon with a wavelength that depends on the type of fluorophore used (typically in the visible spectrum).
Brain slice from a doubly transgenic mouse that expresses CFP and GFP in subsets of cortical neurons.
J Lichtman and B Kasthuri, Washington University School of Medicine, USA
Multicolor Labeling of Cortical Neurons with Lipophilic Dye-Coated Particles.
Fixed brain slice from a P20 mouse that was shot with a combination of the seven different particle types. The image represents a projection of 132 confocal planes covering 50 microns of depth. Many of the cells had processes that extended beyond the upper or lower surface of the slice and hence have dendritic or axonal branches that end abruptly.
Fixed brain slice from a P20 mouse that was shot with a combination of the seven different particle types. The image represents a projection of 132 confocal planes covering 50 microns of depth. Many of the cells had processes that extended beyond the upper or lower surface of the slice and hence have dendritic or axonal branches that end abruptly.
Gan W, et.al
Neuron 27:219-22, 2000.
Jeff W Lichtman,
Washington University School of medicine
St Louis, Missouri, USA
Prospects
for recording from Awake but Head-Restrained Animals (A) Photograph of
a trained rat that is awakeand head-restrained, ready for imaging of
organic [Ca2+]-sensitive dyes. All aspects of the recording procedure
demonstrated in primates are expected hold for mice as well. (Jack
Waters, 2005)