Grafts were cut into 2 sections (cranial and caudal) and each placed in 10% neutral buffered formalin and gently shaken for 48 hours, which allowed the tissue to fix. Each graft sample was then embedded in paraffin wax, sectioned, and stained with hematoxylin and eosin and α-SMA (Sigma-A2547, St. Louis, Mo.) fluorescent dye (for visualization of vascularity, which has been shown to be a measure of graft take).22 Histological sections were analyzed using an Olympus (Center Valley, Pa.) IX51 epifluorescence microscope equipped with an Optronics (Houston, Tex.) Microfire Color CCD Camera and viewed under fluorescence when appropriate. Images of both hematoxylin and eosin and α-SMA sections were viewed and analyzed with National Institutes of Health (Bethesda, Md.) ImageJ software. These analyses were then utilized for statistical comparison.
Microfire color ccd camera
Manufactured by Optronics
Sourced in United States
The Microfire Color CCD Camera is a high-performance laboratory equipment designed for digital image capture and analysis. It features a color CCD sensor that can produce detailed and accurate images. The camera's core function is to convert optical images into digital signals for further processing and analysis.
Automatically generated - may contain errors
Lab products found in correlation
Lsm800 confocal microscope, by Zeiss (2 mentions)
Zen 2.3 software system, by Zeiss (2 mentions)
Stereo investigator software, by MicroBrightField (2 mentions)
C80i microscope, by Nikon (2 mentions)
α sma, by Merck Group (1 mentions)
Dm2000 upright compound microscope, by Leica (1 mentions)
3 3 diaminobenzidine (dab), by Biocare Medical (1 mentions)
Umap anti rb, by Roche (1 mentions)
Dm2000 compound epifluorescence microscope, by Optronics (1 mentions)
6 protocols using microfire color ccd camera
1
Histological Analysis of Graft Vascularity
2
Quantifying Retinal Neurodegeneration in Mice
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Right eyes from deeply anesthetized Herc3-/- and Herc3+/+ mice were collected and processed for histology as described before3 (link). Hematoxylin and Eosin (H&E) staining of retinal sections were prepared for all samples (n = 6 per group). The entire length of H&E retinal sections was imaged at 20 × magnification on both sides of the Optic Nerve Head (ONH) using a Leica DM2000 Upright Compound microscope (Leica Microsystems, Wetzlar, Germany) equipped with an Optronics Microfire color CCD camera (Optronics, Goleta, CA, USA). The H&E images were opened in ImageJ and the ONL thickness was measured at 300 μm intervals starting from the ONH on either side. For each location, we took three measurements within a 20 μm distance from the 300 μm mark and averaged them. The number of layers of nuclei in the ONL was also counted at the same locations by using the duplicate tool in ImageJ. To this end, a rectangle was drawn consistently to one side of the 300 μm mark making sure to include three columns of cells per duplicate image. The number of cells in each of the three columns of nuclear layers was averaged at each 300 μm location. Thus, for each 300 μm location we reported two different parameters: # of nuclei per column of ONL, and ONL thickness in microns.
3
Immunohistochemical Detection of Cleaved Caspase-3
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Immunohistochemistry (IHC) detection of cleaved caspase-3 in formalin-fixed paraffin-embedded (FFPE) tumor tissue was performed according to vendor’s manual instruction (Biocare) and following a verified protocol in the Pathology Laboratory of Translational Medicine at WVU. Briefly, 3 μm sections were deparaffinized on slides, quenched with hydrogen peroxide, and incubated in cleaved caspase-3 antibody (prediluted rabbit polyclonal antibody, Biocare Medical, Concord, CA, 1:200 dilution) at 4 °C for 4 min. Horseradish peroxidase-containing secondary antibody (UMap anti-RB, Roche, Diagnostic, Cupertino, CA) was then added for 8 min and developed using Biocare DAB (brown color). Hematoxylin was used as a counterstain (blue color).
Caspase-3 IHC slides were examined under an Olympus AX70 Provis microscope equipped with an Optronics MicroFire color CCD camera and a 20x/0.70 UPlanApo objective or a 40x/0.90 UApo objective. Images were acquired using Stereo Investigator 10 (MDF Bioscience). Quantification of cells expressing cleaved caspase-3 was performed by ImageJ [25 (link)].
Caspase-3 IHC slides were examined under an Olympus AX70 Provis microscope equipped with an Optronics MicroFire color CCD camera and a 20x/0.70 UPlanApo objective or a 40x/0.90 UApo objective. Images were acquired using Stereo Investigator 10 (MDF Bioscience). Quantification of cells expressing cleaved caspase-3 was performed by ImageJ [25 (link)].
4
Adipose Tissue Morphology Analysis
5
Quantifying Fluorescent Plaque-Associated Markers
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Image acquisition of fluorescent staining images was performed using a Zeiss LSM800 confocal microscope supported by the Zen software (2.3 system) using the × 4 and × 40 lenses, as described previously [25 (link)]. The number of 6E10, Iba-1 associated with plaques, was quantified by unbiased stereological analysis [26 (link)] using the Stereo Investigator software (version 6.02.1, MicroBrightfield) attached to a Nikon C80i microscope equipped with a motorized stage (Ludl) attached to Microfire CCD color camera (Optronics). Four to six sections were analyzed for each animal.
6
Quantification of Amyloid-Beta in Mouse Brain
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Image acquisition of fluorescent staining images was performed using a Zeiss LSM800 confocal microscope supported by the Zen software (2.3 system) using the 4 × and 40 × lenses as described previously (25) . Number of 6E10, Iba-1 associated with plaques were quantified by unbiased stereological analysis (26) using Stereo Investigator software (version 6.02.1, MicroBrightfield) attached to a Nikon C80i microscope equipped with a motorized stage (Ludl) attached to Microfire CCD color camera (Optronics). Four to six sections were analyzed for each animal.
Soluble Aβ 1-42 /Aβ 1-40 ELISA Brain levels of soluble Aβ 1-42 and Aβ 1-40 were quantified by using the Human Amyloid β42 and Human Amyloid β40 Brain ELISA kits (Millipore, Billerica, MA, USA). The experimental procedure was performed according to the manufacturer's instructions (27) .
Soluble Aβ 1-42 /Aβ 1-40 ELISA Brain levels of soluble Aβ 1-42 and Aβ 1-40 were quantified by using the Human Amyloid β42 and Human Amyloid β40 Brain ELISA kits (Millipore, Billerica, MA, USA). The experimental procedure was performed according to the manufacturer's instructions (27) .
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