Live to dead cell imaging.
Live cell imaging allows several key apoptotic events to be visualized in a single cell over time. These include mitochondrial outer membrane permeabilization (MOMP), mitochondrial dysfunction, phosphatidylserine exposure, and membrane permeabilization. Here we describe a protocol for imaging multiple apoptotic processes in the same cell over time. Initially, this involves generating a cell line stably expressing a fluorescent fusion protein that can act as an apoptotic marker, such as cytochrome c-GFP. By combining various fluorescent fusion proteins and probes, several apoptotic events can be imaged in the same cell. Next, the cells are induced to undergo apoptosis and continuously imaged. Finally, quantitative kinetic analysis of various apoptotic processes is performed postimaging.
Tait
Stephen W G
SW
Department of Immunology, St Jude Children's Research Hospital, Memphis, TN, USA.
Bouchier-Hayes
Lisa
L
Oberst
Andrew
A
Connell
Samuel
S
Green
Douglas R
DR
D000818:Animals; D017209:Apoptosis; D051017:Apoptosis Regulatory Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D045304:Cytochromes c / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D003584:Cytological Techniques; D005453:Fluorescence; D006801:Humans; D007700:Kinetics; D018613:Microscopy, Confocal / Q000379:methods*; D008928:Mitochondria / Q000378:metabolism; D024101:Mitochondrial Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism
D051017:Apoptosis Regulatory Proteins; D024101:Mitochondrial Proteins; D011993:Recombinant Fusion Proteins; D045304:Cytochromes c