Transient Room Lighting for Ambient Light Multiphoton Microscopy

Laser scanning microscopy techniques such as confocal and multiphoton fluorescence microscopy have been widely adopted by the biological research community due to their ability to monitor intact specimens at high spatial and temporal resolution. However, they have been limited for many biomedical, clinical and industrial applications by their fundamental need to operate in near absolute darkness. We present a lighting system that allows the use of light-sensitive imaging techniques in a fully-lit room by interleaving capture and illumination at a high frequency and exploiting the light averaging properties of the human eye. We use this system with a multiphoton fluorescence microscope to illustrate that this method is capable of image capture in a well-lit room on par with capture in absolute darkness. This comparison is quantified through noise analysis of the images. This system has been implemented for laser scanning microscopy but has potential for widefield fluorescence imaging suitable for open-field surgery.