Pox, dyes, and videotape: making movies of GFP-labeled vaccinia virus.
The large size of poxvirus virions (approx 250 x 350 nm) makes them ideal candidates for microscopic studies. Recombinant vaccinia viruses that express a viral envelope-specific, green fluorescent protein (GFP) chimera produce enveloped virions that fluoresce green. This fluorescent labeling allows the live, real-time study of viral egress using a variety of microscopic techniques. The methods presented here describe how to image the movement of intracellular enveloped virions that are labeled with green fluorescent protein using time-lapse laser scanning confocal microscopy. Details are also provided for analyzing the images obtained and converting them into QuickTime movies suitable for presentation.
Ward
Brian M
BM
Laboratory of Viral Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, USA.
D002460:Cell Line; D005456:Fluorescent Dyes / Q000378:metabolism; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000378:metabolism*; D008856:Microscopy, Fluorescence / Q000379:methods; D018715:Microscopy, Video / Q000379:methods*; D014616:Vaccinia virus / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D014743:Videotape Recording / Q000379:methods
D005456:Fluorescent Dyes; D008164:Luminescent Proteins; D049452:Green Fluorescent Proteins