Modern approaches for ultrastructural analysis of the zebrafish embryo.
The zebrafish is a powerful vertebrate system with great advantages for both forward and reverse genetic screens and as a model for human disease conditions. Light microscopy has been used extensively to study zebrafish development but less frequently have these studies been combined with ultrastructural information. Zebrafish embryos are ideal for electron microscopy (EM) with a single transverse section containing many different cell types and tissues. However, conventional methods of EM do not provide optimal preservation of all tissues and are usually incompatible with immunolabelling and visualisation of expressed fluorescently tagged proteins. Here we examine methods that overcome these problems. We summarise a range of methods, applicable to the ultrastructural analysis of zebrafish embryos, including methods for fast freezing and processing of zebrafish embryos. These methods preserve antigenicity, ultrastructure and GFP fluorescence even after embedding in resin. In addition, they are compatible with electron tomography. These methods provide a new set of research tools that provide an additional level of information, complementing current methods for study of this widely used model system.
Schieber
Nicole L
NL
Centre for Microscopy and Microanalysis, The University of Queensland, Brisbane, 4072 Queensland, Australia.
Nixon
Susan J
SJ
Webb
Richard I
RI
Oorschot
Viola M J
VM
Parton
Robert G
RG
D000818:Animals; D015925:Cryopreservation / Q000379:methods; D055032:Electron Microscope Tomography / Q000379:methods; D004625:Embryo, Nonmammalian / Q000648:ultrastructure*; D016591:Histocytological Preparation Techniques / Q000379:methods; D006801:Humans; D008854:Microscopy, Electron / Q000295:instrumentation / Q000379:methods*; D008856:Microscopy, Fluorescence / Q000295:instrumentation / Q000379:methods; D015027:Zebrafish / Q000033:anatomy & histology* / Q000196:embryology*