Abstract
Multiphoton microscopy can resolve fluorescent structures and dynamics deep in scattering tissue, but applying this technique in vivo can be limited by short working distance water-immersion objectives. Here we present an ultra long working distance (20 mm) air objective called the Cousa objective. It is optimized for performance across multiphoton imaging wavelengths, offers a > 4 mm2 field-of-view with submicron lateral resolution, and is compatible with commonly used multiphoton imaging systems. We share the full optical prescription, and report performance including in vivo 2-photon and 3-photon imaging in a range of species and preparations.
Competing Interest Statement
The design of the objective is not patented, and it will not be patented in the future. All designs originating in this report are free for reuse, no licensing or material transfer agreements are required. Notification is not required either, but only humbly requested. SLS is a paid consultant for companies that sell optics and multiphoton microscopes. CHY and SLS have interests in the company Pacific Optica. There are no other competing interests.
Footnotes
The revisions are extensive and include new data and analysis. All figures have been revised, many with new panels, and nine additional figures have been added. The text has been revised as well.
