tetano
Editor, Senior Moderator
Methods Mol Biol. 2018;1836:261-279. doi: 10.1007/978-1-4939-8678-1_13.
[h=1]Influenza Virus-Liposome Fusion Studies Using Fluorescence Dequenching and Cryo-electron Tomography.[/h] Gui L[SUP]1,[/SUP][SUP]2[/SUP], Lee KK[SUP]3,[/SUP][SUP]4[/SUP].
[h=3]Author information[/h]
[h=3]Abstract[/h] Influenza virus enters host cells by fusion of viral and endosomal membranes mediated by the influenza hemagglutinin (HA). The pathway of HA-catalyzed fusion has been widely investigated in influenza virus membrane fusion with liposomes. In this chapter we describe methodology for studying the virus-liposome fusion system using a combination of fluorescence dequenching assays and cryo-electron tomography (cryo-ET). In particular, the fluorescence dequenching is used to monitor the efficiency of membrane fusion between whole influenza viruses labeled with a lipophilic dye (DiD) in the membrane and liposomes labeled with a water-soluble dye (sulforhodamine B). By simultaneously monitoring the two fluorescent signals, we can determine the relative time scales of liposomal content leakage or transfer vs. lipid merging. In addition, cryo-ET offers a means of imaging three-dimensional snapshots of different stages of virus-liposome fusion such as prefusion, fusion intermediates, and postfusion.
[h=4]KEYWORDS:[/h] Cryo-electron tomography; Fluorescence spectrometry; Influenza virus; Liposomes; Membrane fusion
PMID: 30151578 DOI: 10.1007/978-1-4939-8678-1_13
[h=1]Influenza Virus-Liposome Fusion Studies Using Fluorescence Dequenching and Cryo-electron Tomography.[/h] Gui L[SUP]1,[/SUP][SUP]2[/SUP], Lee KK[SUP]3,[/SUP][SUP]4[/SUP].
[h=3]Author information[/h]
[h=3]Abstract[/h] Influenza virus enters host cells by fusion of viral and endosomal membranes mediated by the influenza hemagglutinin (HA). The pathway of HA-catalyzed fusion has been widely investigated in influenza virus membrane fusion with liposomes. In this chapter we describe methodology for studying the virus-liposome fusion system using a combination of fluorescence dequenching assays and cryo-electron tomography (cryo-ET). In particular, the fluorescence dequenching is used to monitor the efficiency of membrane fusion between whole influenza viruses labeled with a lipophilic dye (DiD) in the membrane and liposomes labeled with a water-soluble dye (sulforhodamine B). By simultaneously monitoring the two fluorescent signals, we can determine the relative time scales of liposomal content leakage or transfer vs. lipid merging. In addition, cryo-ET offers a means of imaging three-dimensional snapshots of different stages of virus-liposome fusion such as prefusion, fusion intermediates, and postfusion.
[h=4]KEYWORDS:[/h] Cryo-electron tomography; Fluorescence spectrometry; Influenza virus; Liposomes; Membrane fusion
PMID: 30151578 DOI: 10.1007/978-1-4939-8678-1_13