Least invasive in vivo imaging using harmonic generation microscopy

In this manuscript, we review the physics and recent developments of the least invasive optical higher harmonic generation microscopy, with an emphasis on the in vivo molecular imaging applications. Optical higher harmonicgenerations, including second harmonic generation (SHG) and third harmonic generation (THG), leave no energy deposition to the interacted matters due to their energy-conservation characteristic, providing the "noninvasiveness" nature desirable for clinical studies. Combined with their nonlinearity, harmonic generation microscopy provides threedimensional sectioning capability, offering new insights into live samples. By choosing the lasers working in the high penetration window, we have recently developed a least-invasive in vivo light microscopy with submicron 3D resolution and high penetration, utilizing endogenous and resonantly-enhanced multi-harmonic-generation signals in live specimens, with focused applications on the developmental biology study and clinical virtual biopsy.