2-Photon microscopes: detectors and collection efficiency
All detectors are GaAs photomultiplier tubes hand selected for highest quantum efficiency.
The microscopes can accept up to four detectors: two of them are collecting light coming through the objective, the two others are collecting fluorescent light coming out of the condenser. The signals from both paths can be added for higher signal to noise ratio.
While collection from the condenser does not represent a major challenge, scattering in the sample makes the light collected through the objective more difficult to redirect toward the detector. We have designed a fluorescence collection system that nearly maximize the collection efficiency. A carefully selected set of lenses, and a reduced path length result in a short detector assembly mounted in close proximity to the objective lens.
2-Photon microscopes: speed
The maximum speed of our microscopes is 0.5ms per line, obtained in bi-directional scanning. For a typical 256*256pixels image that corresponds to 128ms per frame, or ~8 frames per seconds.
Some applications can accommodate acquisitions of images with fewer lines, reducing proportionally the acquisition time, and increasing the frame rate. For example, we routinely acquire 256*96 z-stacks at 18 frames per seconds.
A number to remember: To form an image with sufficient signal to noise ratio, a sufficient number of photons need to be collected per pixel. On a typical biological sample, with typical power used to preserve samples from rapid damage, one needs to spend a few microseconds on each pixel to collect enough photons. Consequently, speeds exceeding .5ms per line are rarely of any use.