In vivo developmental biology study using noninvasive multi-harmonic generation microscopy

Morphological changes and complex developmental processes inside vertebrate embryos are difficult to observe noninvasively with millimeter-penetration and sub-micrometer-resolution at the same time. By using higher harmonic generation, including second and third harmonics, as the microscopic contrast mechanism, optical noninvasiveness can be achieved due to the virtual-level-transition characteristic. The intrinsic nonlinearity of harmonic generations provides optical sectioning capability while the selected 1230-nm near-infrared light source provides the deep-penetration ability. The complicated development within a similar to 1.5-mm thick zebrafish (Danio rerio) embryo from initial cell proliferation, gastrulation, to tissue formation can all be observed clearly in vivo without any treatment on the live specimen. (C) 2003 Optical Society of America.