Digitizing life at the level of the cell: high-performance laser-scanning microscopy and image analysis for in toto imaging of development.
The field of biological imaging is progressing at an amazing rate. Advances in both laser-scanning microscopy and green fluorescent protein (GFP) technology are combining to make possible imaging-based approaches for studying developmental mechanisms that were previously impossible. Modern confocal and multi-photon microscopes are pushing the envelope of speed, sensitivity, spectral resolution, and depth resolution to allow in vivo imaging of whole, live embryos at cellular resolution over extended periods of time. In toto imaging, in which nearly every cell in an embryo or tissue can be tracked through space and time during development, may become a standard technique for small transparent embryos such as zebrafish and early stage chick and mouse embryos. GFP and its spectral variants can be used to mark a wide range of in vivo biological information for in toto imaging including gene expression patterns, mutant phenotypes, and protein subcellular localization patterns. Combining in toto imaging and GFP transgenic approaches on a large scale may usher in an explosion of in vivo, developmental data as has happened in the past several years with genomic data. There are significant challenges that must be met to reach these goals. This paper will discuss the current state-of-the-art, the challenges, and the prospects of in toto imaging in the areas of imaging, image analysis, and informatics.
Megason
Sean G
SG
Biological Imaging Center, Beckman Institute and Division of Biology, California Institute of Technology, Pasadena, CA 91125, USA. megason@caltech.edu
Fraser
Scott E
SE
D000818:Animals; D030801:Animals, Genetically Modified; D002642:Chick Embryo; D018507:Gene Expression Regulation, Developmental; D049452:Green Fluorescent Proteins; D007091:Image Processing, Computer-Assisted / Q000379:methods*; D008164:Luminescent Proteins / Q000378:metabolism; D051379:Mice; D018613:Microscopy, Confocal / Q000379:methods*; D008856:Microscopy, Fluorescence; D017785:Photons; D012984:Software; D013997:Time Factors; D015027:Zebrafish
D008164:Luminescent Proteins; D049452:Green Fluorescent Proteins