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Here, we provide detailed protocols for building an experimental system that employs atomic force microscopy and a single-molecule DNA tightrope assay based on oblique angle illumination fluorescence microscopy.
Research began by using various non-invasive analyses (photographs under ultraviolet illumination, fluorescence lifetime imaging, x-ray fluorescence).
During illumination, fluorescence measurements and light dosimetry were performed.
Photobleaching was prevented by shuttering the illumination fluorescence pathway with a filter wheel.
Images of the stained gels were captured with the Biorad 4.2.1 Fluor-S MultiImager system (Biorad) using top illumination fluorescence.
The structured illumination fluorescence image, on the other hand, clearly shows axial distribution of the blood vessels, shown in Fig. 2(b).
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Sections were examined using an epi-illumination fluorescence microscope (Zeiss Axioskop 2).
Alexa 488 or 594-conjugated IgG (Invitrogen) was used as a secondary antibody, and stained cells were observed by epi-illumination fluorescence microscope (Olympus High Content microscopy).
121, 133– 137 An optical imaging technique commonly used is two-dimensional (2D) fluorescence reflectance imaging (FRI), also known as epi-illumination fluorescence imaging.
The optical tweezers set-up (Fig. 1) is based on an inverted microscope (Axiovert S 100 TV, Carl Zeiss, Jena, Germany) which is designed for epi-illumination fluorescence microscopy.
During the experiment, the real time image of iris blood vessel was recorded by an epi-illumination fluorescence video microscopy system (Optiphot 2, Nikon, Japan) equipped with a 100 W mercury lamp, CCD camera (Hamamatsu C2400, Japan), a video recorder (VC-S5, Sharp, Japan) with a video timer (VTG-33, For-A, Japan), and 20x objective lens (CF Plan Fluor, Nikon, Japan).
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