Scanning surface confocal microscopy for simultaneous topographical and fluorescence imaging: application to single virus-like particle entry into a cell.
We have developed a method for simultaneous recording of high-resolution topography and cell surface fluorescence in a single scan which we call scanning surface confocal microscopy. The resolution of the system allows imaging of individual fluorescent particles in the nanometer range on fixed or live cells. We used this technique to record the interaction of single virus-like particles with the cell surface and demonstrated that single particles sink into the membrane in invaginations reminiscent of caveolae or pinocytic vesicles. This method provides a technique for elucidating the interaction of individual viruses and other nanoparticles, such as gene therapy vectors, with target cells. Furthermore, this technique should find widespread application for studying the relationship of fluorescently tagged molecules with components of the cell plasma membrane.
Gorelik
J
J
Division of Medicine and Medical Research Council Clinical Sciences Centre, Imperial College London, Hammersmith Hospital Campus, Du Cane Road, London W12 0NN, United Kingdom.
Shevchuk
A
A
Ramalho
M
M
Elliott
M
M
Lei
C
C
Higgins
C F
CF
Lab
Max J
MJ
Klenerman
D
D
Krauzewicz
N
N
Korchev
Y
Y
D000818:Animals; D019556:COS Cells / Q000648:ultrastructure / Q000821:virology*; D002213:Capsid / Q000502:physiology / Q000648:ultrastructure*; D002232:Carbocyanines; D002462:Cell Membrane / Q000648:ultrastructure / Q000821:virology; D002522:Chlorocebus aethiops; D004867:Equipment Design; D005456:Fluorescent Dyes; D005813:Genes, Synthetic; D018613:Microscopy, Confocal / Q000295:instrumentation / Q000379:methods*; D008856:Microscopy, Fluorescence / Q000295:instrumentation / Q000379:methods*; D036103:Nanotechnology / Q000379:methods*; D010316:Particle Size; D013499:Surface Properties
D002232:Carbocyanines; D005456:Fluorescent Dyes; C089187:cyanine dye 3