Performing confocal microscopy with two microscope objectives on either side of the sample increases the axial resolution for improved three-dimensional imaging.
A suture was tied around the distal tendon, which was then separated, and the muscle was reflected over the microscope objective of a Nikon Diaphot 300 inverted microscope (Nikon Canada, Mississauga, ON, Canada) with the proximal neurovascular bundle intact.
While image quality is decreased slightly when compared to a microscope objective of similar NA, many of the important tissue features are identifiable, show similarity to anatomic features seen in histology slides, and are potentially useful for diagnosis [ 27].
To activate ChR2-expressing fibers, light from a 473-nm laser (Optoengine, Midvale, UT) was focused on the back aperture of the microscope objective to produce wide-field illumination of the recorded cell.
It was precisely aligned by moving the translation stage in the XYZ directions to guide the transmitted light from the microscope objective to the center of the fiber.
The short depth of focus of optical microscope objectives allows some kind of vertical multiplexing of a biological image with a PRP image.
This method utilizes the full depth of field of the microscope objectives focusing different planes of the thick cortex.
In order to interpret our interference data, we have assumed that the electric field collected by the microscope objective consists of a reflection of the incident electric field from the coverslip-buffer interface and an additional, single reflection from a discrete, protein dense, site near the ventral plasma membrane.
Future improvements of cell tracking techniques in vivo requires, among others, the development of specifically designed new fluorophores with improved two-photon absorption cross-sections and the optimization of microscope objectives and detector light paths to maximize the collection of emitted fluorescence photons.
When imaging the reflection from a tilt-free, planar reflector at the focus of a high numerical microscope objective, all of the incident rays will be collected by the objective, however, for a sphere the angle of reflection of an incident ray at the sphere's surface depends upon the transverse scan position.
