Fluorescence imaging and spectroscopy of biomaterials in air and liquid by scanning near-field optical/atomic force microscopy.
We have developed scanning near-field optical/atomic force microscopy (SNOM/AFM). The SNOM/AFM uses a bent optical fiber simultaneously as a dynamic force AFM cantilever and a SNOM probe. Resonant frequency of the optical fiber cantilever is 15-40 kHz. Optical resolution of the SNOM/AFM images shows less than 50 nm. The SNOM/AFM system contains photon counting system and polychrometer/intensified coupled charge devise (ICCD) system to observe fluorescence image and spectrograph of micro areas, respectively. Cultured cells were stained with fluorescein isothiocyanate (FITC)-labeled anti-keratin antibody or FITC-labeled phalloidin after treatment with Triton X-100. Fluorescence and topographic images were obtained in air and water. The fluorescence images showed clear images of keratin and actin filaments. The SNOM/AFM is perfect to observe biomaterials in liquid with a liquid chamber while the topographic Images showed subcellular structures which correspond to keratin and actin filaments.
Muramatsu
H
H
Research Lab. for Advanced Tech., Seiko Instruments Inc., Chiba, Japan. muramatu@tk.sii.co.jp
Chiba
N
N
Nakajima
K
K
Ataka
T
T
Fujihira
M
M
Hitomi
J
J
Ushiki
T
T
D000199:Actins / Q000032:analysis; D000388:Air; D002294:Carcinoma, Squamous Cell; D002857:Chromium / Q000737:chemistry; D003599:Cytoskeleton / Q000648:ultrastructure*; D004938:Esophageal Neoplasms; D019793:Fluorescein; D006801:Humans; D007633:Keratins / Q000032:analysis; D018625:Microscopy, Atomic Force / Q000295:instrumentation / Q000379:methods; D008855:Microscopy, Electron, Scanning / Q000295:instrumentation / Q000379:methods; D013050:Spectrometry, Fluorescence / Q000379:methods*; D014407:Tumor Cells, Cultured; D014867:Water
D000199:Actins; D014867:Water; D002857:Chromium; D007633:Keratins; D019793:Fluorescein