Two-photon excitation microscopy is a fluorescence imaging technique that is particularly well-suited to image scattering living tissue of up to about one millimeter in thickness. Unlike traditional f
Two-photon excitation microscopy of mouse intestine. Red: actin. Green: cell nuclei. Blue: mucus of goblet cells. Obtained at 780 nm using a Ti-sapphire laser.
Two-photon fluorescence image of a stained cross section of a lily of the valley rhizome. The excitement is at 840 nm, and the red and blue colors represent other channels of multiphoton techniques which have been superimposed.
Schematic representation of the energy levels (Jabłoński diagrams) of the fluorescence process, example of a fluorescent dye that emits light at 460 nm. One (purple, 1PEF), two (light red, 2PEF) or three (dark red, 3PEF) photons are absorbed to emit a photon of fluorescence (turquoise).
Optical response from a point source. From left to right: calculated intensity distributions xy (top) and rz (bottom), with logarithmic scale, for a point source imaged by means of a wide field (a), 2PEF (b) and confocal microscopy (c). The 2PEF and confocal forms have a better signal-to-noise ratio than the wide field. The 2PEF distribution is larger due to the fact that a wavelength twice as long as in the case of a wide or confocal field is responsible for the intensity distribution. These intensity distributions are also known as point spread functions. Optical conditions: the excitation wavelengths are 488 nm and 900 nm respectively for 1PEF and 2PEF; the emission wavelength is 520 nm; the
Fluorescence microscope
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A fluorescence microscope is an optical microscope that uses fluorescence instead of, or in addition to scattering, reflection, and attenuation or absorption, to study the properties of organic or ino
Two-photon excitation microscopy
…imaging technique that is particularly well-suited to image scattering living tissue of up to about one millimeter in thickness. Unlike traditional fluorescence microscopy, where the excitation wavelength is shorter than the emission wavelength, two-photon excitation requires simultaneous excitation by two photons with…
An upright fluorescence microscope (Olympus BX61) with the fluorescence filter cube turret above the objective lenses, coupled with a digital camera
A sample of herring sperm stained with SYBR green in a cuvette illuminated by blue light in an epifluorescence microscope. The SYBR green in the sample binds to the herring sperm DNA and, once bound, fluoresces giving off green light when illuminated by blue light.
Epifluorescent imaging of the three components in a dividing human cancer cell. DNA is stained blue, a protein called INCENP is green, and the microtubules are red. Each fluorophore is imaged separately using a different combination of excitation and emission filters, and the images are captured sequentially using a digital CCD camera, then superimposed to give a complete image.
Endothelial cells under the microscope. Nuclei are stained blue with DAPI, microtubules are marked green by an antibody bound to FITC and actin filaments are labeled red with phalloidin bound to TRITC. Bovine pulmonary artery endothelial (BPAE) cells