title: Continuous wave two-photon scanning near-field optical microscopy.
abstract: We have implemented continuous-wave two-photon excitation of near-UV absorbing fluorophores in a scanning near-field optical microscope (SNOM). The 647-nm emission of an Ar-Kr mixed gas laser was used to excite the UV-absorbing DNA dyes DAPI, the bisbenzimidazole Hoechst 33342, and ethidium bromide in a shared aperture SNOM with uncoated fiber tips. Polytene chromosomes of Drosophila melanogaster and the nuclei of 3T3 Balb/c cells labeled with these dyes were readily imaged. The fluorescence intensity showed the expected nonlinear (second order) dependence on the excitation power in the range of 8-180 mW. We measured the fluorescence intensity as a function of the tip-sample displacement in the direction normal to the sample surface in the single- and two-photon excitation modes (SPE, TPE). The fluorescence intensity decayed faster in TPE than in SPE.
authors.affiliation: Department of Molecular Biology, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany.
authors.forename: A K
authors.initials: AK
authors.lastname: Kirsch
authors.forename: V
authors.initials: V
authors.lastname: Subramaniam
authors.forename: G
authors.initials: G
authors.lastname: Striker
authors.forename: C
authors.initials: C
authors.lastname: Schnetter
authors.forename: D J
authors.initials: DJ
authors.lastname: Arndt-Jovin
authors.forename: T M
authors.initials: TM
authors.lastname: Jovin
mesh_terms: D016475:3T3 Cells; D000818:Animals; D055592:Biophysical Phenomena; D001703:Biophysics; D002467:Cell Nucleus / Q000648:ultrastructure; D002875:Chromosomes / Q000648:ultrastructure; D004331:Drosophila melanogaster; D005456:Fluorescent Dyes; D007211:Indoles; D051379:Mice; D008856:Microscopy, Fluorescence / Q000295:instrumentation / Q000379:methods*; D017785:Photons
chemical_list: D005456:Fluorescent Dyes; D007211:Indoles; C007293:DAPI