Transfected single-cell imaging by scanning electrochemical optical microscopy with shear force feedback regulation.
Gene-transfected single HeLa cells were characterized using a scanning electrochemical/optical microscope (SECM/OM) system with shear-force-based probe-sample distance regulation to simultaneously capture electrochemical, fluorescent, and topographic images. The outer and inner states of single living cells were obtained as electrochemical and fluorescent signals, respectively, by using an optical fiber-nanoelectrode probe. A focused ion beam (FIB) was used to mill the optical aperture and the ring electrode at the probe apex (the inner and outer radii of the ring electrode were 37 and 112 nm, respectively). To apply an appropriate shear force between the probe tip and the living cell surface, we optimized the amplitude of oscillation of the tuning fork to which the probe was attached. Field-programmable gate arrays (FPGA) were adopted to drastically increase the feedback speed of the tip-sample distance regulation, shorten the scanning time for imaging, and enhance the accuracy and quality of the living cell images. In employing these improvements, we simultaneously measured the cellular expression activity of both secreted alkaline phosphatase outside and GFP inside by using the SECM/OM with shear force distance regulation.
Takahashi
Yasufumi
Y
Graduate School of Environmental Studies, Tohoku University, Aramaki Aoba 6-6-11-605, Sendai 980-8579, Japan.
Shiku
Hitoshi
H
Murata
Tatsuya
T
Yasukawa
Tomoyuki
T
Matsue
Tomokazu
T
D000469:Alkaline Phosphatase / Q000235:genetics / Q000378:metabolism; D000814:Aniline Compounds / Q000378:metabolism; D002470:Cell Survival; D004563:Electrochemistry; D004566:Electrodes; D005110:Extracellular Space / Q000378:metabolism; D005822:Genetic Vectors / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D008853:Microscopy; D057054:Molecular Imaging / Q000379:methods*; D055100:Optical Fibers; D055620:Optical Phenomena; D009943:Organophosphorus Compounds / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D014162:Transfection
D000814:Aniline Compounds; D009943:Organophosphorus Compounds; D049452:Green Fluorescent Proteins; C068798:4-aminophenylphosphate; D000469:Alkaline Phosphatase