Freely tunable spectral detection for multiphoton microscopy

摘要

Multiphoton microscopy enables imaging deep inside living specimens. High photon flux is required for the nonlinearexcitation of fluorescent markers, which confines the excitation to the small volume of the focal spot. This results inintrinsic optical sectioning and enables non-descanned detection of the fluorescence signal. Although tunable spectraldetection has been standard in confocal microscopy for many years, it is still common in multiphoton microscopy tomanually change the filters in the detection beam path of the microscope to accommodate for different fluorescencemarkers. We revolutionized the non-descanned detection by implementing a freely tunable spectral detector for fourdetection channels. This enables the adaptation to new transgenic markers in seconds and separates strongly overlappingspectra without mathematical restoration. Here, we introduce the technological concept and show its application inimaging of biological specimens demonstrating the capability of the spectral detector.