Eric Betzig, a scientist from Janelia Farm Research Campus at the Howard Hughes Medical Institute, in collaboration with Na Ji, a postdoctoral scientist, has enhanced live brain imaging by integrating two-photon fluorescence microscopy with adaptive optics. The technique can aid researchers to view the mouse brain more clearly.
Astronomers generally use adaptive optics to define the celestial body images. They direct a laser beam in the atmosphere typically in the same direction as the celestial object they want to observe. The light, which is reflected back from the object, gets altered while traveling through the atmosphere. To overcome this problem, astronomers use a wavefront sensor to determine the alteration and then utilize these calculations to change a telescope mirror to stop the atmospheric distortions. This rectification gives a clearer vision of the object.
However, a wavefront sensor cannot be placed directly in a live animal to determine the light distortions inside the tissue. In order to solve this problem, Betzig said that if the rays are examined separately, then their deflections can be rectified and can be directed back to a single point.
Betzig and Ji inserted fluorescent beads into the mouse brain. The beads functioned as navigators to determine the rays’ deflections. To achieve this, the scientists utilized a 1” spatial light modulator, which enabled them to activate individual rays at a time and then capture the head image. The amount of the deflected ray was then determined from the bead's image shift relative to the required focal spot. The display was then used as a compact mirror to direct the ray back to the focal spot. The procedure was again repeated with other rays. This technique enhances both the signal and the optimal resolution.
In another experiment, Ji injected fluorescent bead guide stars and fluorescent neuron marker to mark the brains of mouse fetuses. After the animal matured, the scientist excised a portion of the skull and substituted it with a transparent glass cover. The adaptive optical microscope was then used to capture the neurons via the glass.