Multi-Photon-Laser-Scanning-Microscopy (MPLSM) now stands as one of the most powerful experimental tools in
biology. Specifically, MPLSM based in-vivo studies of structures and processes in the brains of small rodents and
imaging in brain-slices have led to considerable progress in the field of neuroscience.
Equipment allowing for independent control of two laser-beams, one for imaging and one for photochemical
manipulation, strongly enhances any MPLSM platform. Some industrial MPLSM producers have introduced double
scanner options in MPLSM systems.
Here, we describe the upgrade of a single scanner MPLSM system with equipment that is suitable for independently
controlling the beams of two Titanium Sapphire lasers. The upgrade is compatible with any actual MPLSM system and
can be combined with any commercial or self assembled system.
Making use of the pixel-clock, frame-active and line-active signals provided by the scanner-electronics of the MPLSM,
the user can, by means of an external unit, select individual pixels or rectangular ROIs within the field of view of an
overview-scan to be exposed, or not exposed, to the beam(s) of one or two lasers during subsequent scans. The switching
processes of the laser-beams during the subsequent scans are performed by means of Electro-Optical-Modulators
While this system does not provide the flexibility of two-scanner modules, it strongly enhances the experimental
possibilities of one-scanner systems provided a second laser and two independent EOMs are available.
Even multi-scanner-systems can profit from this development, which can be used to independently control any number
of laser beams.