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Signal detection was achieved either by an Axiocam camera connected to an Axioplan2 fluorescence microscope (Zeiss), or by confocal imaging using a TCS SP2 AOBS microscope (Leica).
Increased vascularity was confirmed by confocal imaging using the lectin UEA-1.
Intracellular Ca2+ increase following ATP stimulation was further confirmed by confocal imaging using a different Ca2+-sensitive dye (Fluo-4).
Cytoplasmic calcium levels were measured in BEAS-2B cells by confocal imaging using Fluo-4AM calcium-sensitive dye.
Analysis of the cells was done by confocal imaging using a Leica SP2 AOBS system (Leica Microsystems, Milan, Italy) and a 63 × oil-immersion lens.
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Recording of the cell membrane potential was synchronized with confocal imaging using a TTL synchronizing pulse generated by the confocal scanner at the beginning of the time series protocol.
Confocal imaging using the down tapered DCFC was demonstrated on a spectrally encoded confocal microscopy setup.
We found that ELuc localized to peroxisomes and nuclei, which was verified by confocal fluorescence imaging using peroxisome- and nucleus-targeted GFP (Figure S6), as reported for FLuc [23], [24].
To confirm the scaling effects we observed by confocal imaging and surface biotinylation assays, we used hippocampal slice electrophysiology to measure AMPAR-mediated excitatory postsynaptic responses (EPSCs) in CA1 neurons.
Slides were examined by confocal imaging on a Nikon CS 2, original magnification × 40 (WD 0.24 mm oil) using the NIS software (Nikon Instruments Inc., Melville, NY, USA).
Control experiments using Lysotracker Green (Molecular Probes, 1∶1000 dilution for 30 mins at 37°C) first verified that chloroquine diphosphate treatment disrupted the lysosomes by confocal imaging.
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