Interferometric fluorescent super-resolution microscopy resolves 3D cellular ultrastructure.
Understanding molecular-scale architecture of cells requires determination of 3D locations of specific proteins with accuracy matching their nanometer-length scale. Existing electron and light microscopy techniques are limited either in molecular specificity or resolution. Here, we introduce interferometric photoactivated localization microscopy (iPALM), the combination of photoactivated localization microscopy with single-photon, simultaneous multiphase interferometry that provides sub-20-nm 3D protein localization with optimal molecular specificity. We demonstrate measurement of the 25-nm microtubule diameter, resolve the dorsal and ventral plasma membranes, and visualize the arrangement of integrin receptors within endoplasmic reticulum and adhesion complexes, 3D protein organization previously resolved only by electron microscopy. iPALM thus closes the gap between electron tomography and light microscopy, enabling both molecular specification and resolution of cellular nanoarchitecture.
Shtengel
Gleb
G
Howard Hughes Medical Institute, Janelia Farm Research Campus, Ashburn, VA 20147, USA.
Galbraith
James A
JA
Galbraith
Catherine G
CG
Lippincott-Schwartz
Jennifer
J
Gillette
Jennifer M
JM
Manley
Suliana
S
Sougrat
Rachid
R
Waterman
Clare M
CM
Kanchanawong
Pakorn
P
Davidson
Michael W
MW
Fetter
Richard D
RD
Hess
Harald F
HF
D000818:Animals; D002460:Cell Line; D002522:Chlorocebus aethiops; D006801:Humans; D021621:Imaging, Three-Dimensional / Q000295:instrumentation* / Q000379:methods*; D008856:Microscopy, Fluorescence / Q000295:instrumentation* / Q000379:methods*; D008857:Microscopy, Interference / Q000295:instrumentation* / Q000379:methods*; D008870:Microtubules