Near-field optical and shear-force microscopy of single fluorophores and DNA molecules.
Photodynamics of individual fluorescence molecules has been studied using an aperture-type near-field scanning optical microscope with two channel fluorescence polarisation detection and tuning fork shear-force feedback. The position of maximum fluorescence from individual molecules could be localised with an accuracy of 1 nm. Dynamic processes such as translational and rotational diffusion were observed for molecules adsorbed to a glass surface or embedded in a polymer host. The in-plane molecular dipole orientation could be determined by monitoring the relative contribution of the fluorescence signal in the two perpendicular polarised directions. Rotational dynamics was investigated on 10 ms-1000 s timescale. Shear-force phase feedback was used to obtain topographic imaging of DNA fragments, with a lateral and vertical resolution comparable to scanning force microscopy. A DNA height of 1.4 nm has been measured, an indication of the non-disturbing character of the shear force mechanism.
Garcia-Parajo
M F
MF
Department of Applied Physics, University of Twente, AE Enschede, The Netherlands.
Veerman
J A
JA
Ruiter
A G
AG
van Hulst
N F
NF
D004247:DNA / Q000648:ultrastructure*; D005454:Fluorescence Polarization; D008853:Microscopy / Q000295:instrumentation* / Q000379:methods*; D008855:Microscopy, Electron, Scanning; D008856:Microscopy, Fluorescence / Q000295:instrumentation / Q000379:methods*; D017785:Photons; D012680:Sensitivity and Specificity
D004247:DNA