Ultrastructural in situ hybridization: a review of technical aspects.
Detection of nucleic acid sequence at the ultrastructural level has allowed us to better understand the expression of genes in some fields of application in cell biology. In situ hybridization at the ultrastructural level can be carried out using three different methods: on vibratome sections before embedding in epoxy resin, on ultrathin frozen section, or on ultrathin section of tissue embedded in hydrophilic resin such as Lowicryl. Before starting the detection of nucleic acid sequences at the electron microscope level, the experimenter has to choose various parameters; the type of tissue fixation, the probe and its label, and the in situ hybridization method, depending on the sensitivity, the resolution and the ultrastructural preservation required. This review of technical aspects, by describing the different methods of ultrastructural in situ hybridization, will help the experimenter to optimize each step of the hybridization procedure.
Le Guellec
D
D
Institut de Biologie et Chimie des Protéines, Lyon, France.
D000818:Animals; D001921:Brain / Q000378:metabolism / Q000648:ultrastructure; D002642:Chick Embryo; D003094:Collagen / Q000235:genetics; D005260:Female; D005353:Fibronectins / Q000235:genetics; D005629:Frozen Sections; D015870:Gene Expression; D013006:Growth Hormone / Q000235:genetics; D006801:Humans; D017403:In Situ Hybridization / Q000379:methods*; D008854:Microscopy, Electron / Q000379:methods*; D015335:Molecular Probes; D010121:Oxytocin / Q000235:genetics; D010902:Pituitary Gland / Q000378:metabolism / Q000648:ultrastructure; D012313:RNA / Q000235:genetics / Q000378:metabolism; D051381:Rats; D016610:Tissue Embedding; D016707:Tissue Fixation
D005353:Fibronectins; D015335:Molecular Probes; D010121:Oxytocin; D012313:RNA; D013006:Growth Hormone; D003094:Collagen