Intravital imaging techniques, while powerful, are challenged by major tradeoffs between spatial resolution, the rapidity of data collection and the limited photon budget. This combination of requirements is challenging, but necessary: intravital imaging can only generate accurate data on cell lineages and cell migration if it can capture the three dimensional image of the entire embryo or tissue before any of the cells can move half of the distance separating neighboring cells.
We are attempting to advance these tradeoffs by constructing faster and more efficient microscopes that maintain subcellular resolution. Our new microscope combines the deep penetration of two-photon microscopy and the speed of light sheet microscopy (often termed SPIM) to generate images with more than ten-fold improved imaging speed and sensitivity. This two-photon SPIM is far less subject to light scattering, permitting subcellular resolution to be maintained far better than conventional light sheet microscopes.
Parallel developments in novel labels and new means to interpret the fluctuations in the signals are providing new windows into the molecular signals, their motions and the responses they elicit. Combined, these approaches permit the orchestrated cellular events that build embryos to be interrogated directly to single cell resolution, offering new insights into the events that build embryos and organs, and suggesting novel means to execute cell therapies.