Endocytic pathways: combined scanning ion conductance and surface confocal microscopy study.
We introduce a novel high resolution scanning surface confocal microscopy technique that enables imaging of endocytic pits in apical membranes of live cells for the first time. The improved topographical resolution of the microscope together with simultaneous fluorescence confocal detection produces pairs of images of cell surfaces sufficient to identify single endocytic pits. Whilst the precise position and size of the pit is detected by the ion conductance microscope, the molecular nature of the pit, e.g. clathrin coated or caveolae, is determined by the corresponding green fluorescent protein fluorescence. Also, for the first time, we showed that flotillin 1 and 2 can be found co-localising with approximately 200-nm indentations in the cell membrane that supports involvement of this protein in endocytosis.
Shevchuk
Andrew I
AI
MRC Clinical Sciences Centre, Faculty of Medicine, Imperial College London, Hammersmith Hospital Campus, Du Cane Road, London W12 0NN, UK.
Hobson
Phil
P
Lab
Max J
MJ
Klenerman
David
D
Krauzewicz
Nina
N
Korchev
Yuri E
YE
D000818:Animals; D019556:COS Cells; D021941:Caveolae / Q000648:ultrastructure*; D002462:Cell Membrane / Q000378:metabolism / Q000648:ultrastructure; D002522:Chlorocebus aethiops; D004705:Endocytosis / Q000502:physiology*; D049452:Green Fluorescent Proteins; D007668:Kidney / Q000166:cytology / Q000648:ultrastructure; D008565:Membrane Proteins / Q000378:metabolism / Q000648:ultrastructure; D018613:Microscopy, Confocal / Q000379:methods*; D020527:Microscopy, Scanning Probe / Q000379:methods*
D008565:Membrane Proteins; C106354:flotillins; D049452:Green Fluorescent Proteins