Virtual Histology of Transgenic Mouse Embryos for High-Throughput Phenotyping

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Abstract
A bold new effort to disrupt every gene in the mouse genome necessitates systematic, interdisciplinary approaches to analyzing patterning defects in the mouse embryo. We present a novel, rapid, and inexpensive method for obtaining high-resolution virtual histology for phenotypic assessment of mouse embryos. Using osmium tetroxide to differentially stain tissues followed by volumetric X-ray computed tomography to image whole embryos, isometric resolutions of 27 μm or 8 μm were achieved with scan times of 2 h or 12 h, respectively, using mid-gestation E9.5–E12.5 embryos. The datasets generated by this method are immediately amenable to state-of-the-art computational methods of organ patterning analysis. This technique to assess embryo anatomy represents a significant improvement in resolution, time, and expense for the quantitative, three-dimensional analysis of developmental patterning defects attributed to genetically engineered mutations and chemically induced embryotoxicity. Developmental biology is entering the digital age, thanks to advancements in imaging technologies, instrumentation, and software. These advancements are converging with discoveries in developmental biology to deliver unprecedented insight into how human development is impacted by the products we use, the environment we live in, and our genetic composition. Industrial societies are becoming increasingly concerned with the exposure of women and their unborn fetuses to pharmaceuticals and commonplace household chemicals. In addition, understanding genetic causes of birth defects is now possible through the isolation of specific genes, which can be efficiently disrupted in embryos, and subsequently observed for birth defects. Such studies of embryotoxicity typically involve the use of mouse embryos. However, evaluation of mouse embryos in the past has involved expensive and cumbersome external inspection and thin sectioning for view under the microscope. As such, developmental biologists have eagerly anticipated the advent of tools that would allow them to routinely assess the complex and dynamic organization of embryos using techniques that are fast and inexpensive. In this article, the authors introduce a rapid, high-quality, and inexpensive technique for the three-dimensional visualization of mouse embryos using X-ray computed tomography that is ideally suited for researchers in pharmaceutical, industrial, and academic laboratories.