Label-free multiphoton hyperspectral imaging (Conference Presentation)

We present a novel detection scheme that allows the nonlinear microscope to register the spectral fingerprints of native biomolecules. This tailored instrument leverages a grating to disperse the nonlinear signals, a galvanometric mirror to scan each spectral component, and a high dynamic range photomultiplier tube to detect them. This device co-registers the spectral profiles of each probed sample point, delivering spectra with harmonic and multiphoton absorption fluorescence signals three orders of magnitude faster than a benchmark CCD-based spectrometer. By virtue of spectra, our approach exposes the manifold of biochemical species within the interaction volume, enhancing the granularity needed for investigating biological specimens. This detection scheme offers to nonlinear microscopy a contrast palette with complementary signals and superior analytical power.