ASL volumes from NHBE cells were estimated by meniscus scanning as previously described [20 (link)]. Briefly, cultures were washed with phosphate buffered saline (PBS) 24 h prior to measurements to clean the filters from mucus accumulation, which can interfere with ASL reading. At the time of measurement, the 12 mm Transwell supports were placed on a commercially available scanner (Epson perfection V500 PHOTO, Epson America Inc., Long Beach, CA). Scanned menisci were used to estimate ASL volume using software generously provided by Dr. Myerburg (University of Pittsburgh). Calibration of the 12 mm Transwell supports (Costar Corning, Tewksbury, MA, USA) was done by measuring ASL changes after apical addition of different volumes of PBS.
Perfection v500 photo
Manufactured by Epson
Sourced in Japan, United States, Germany
The Epson Perfection V500 PHOTO is a flatbed scanner designed for high-quality image scanning. It features a maximum optical resolution of 6400 dpi and can scan photographs, slides, and film negatives up to 8.5 x 11.7 inches in size.
Automatically generated - may contain errors
6 protocols using perfection v500 photo
1
ASL Volume Estimation from NHBE Cells
2
Quantifying Radiation-Induced Cell Colonies
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HPV-negative and -positive cells were seeded in 6-well plates at a density comprised between 250 and 2,000 cells per well, depending on their growth rate. After 24h, cells were irradiated with 0, 1, 2, 4 or 6 Gy using a Gammacell 40 Exactor (Best Theratronics) and incubated for 10 days at 37 °C in a humidified 5% CO2 atmosphere. Plates were then washed in PBS, fixed and colored for 45 min using a solution containing 5% glutaraldehyde (Acros Organics, Thermo Fischer scientific) and 0,5% Crystal violet (Sigma-Aldrich, Saint Louis, MI, USA). After a washing step with deionized water, plates were dried, scanned (Epson Perfection V500 PHOTO, Epson, Nagano, Japan) and quantified using the ColonyArea ImageJ plugin (ImageJ software, National Institute of Health, Bethesda, MD, USA). Instead of the traditional manual counting, this standardized/computerized approach allows to determine the percentage of area covered by cell colonies 33 (link).
3
Measuring Cookie Color Using Computer Vision
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The color of the cookies was measured using computer vision as previously described in detail [25 (link)]. Samples were captured with a flatbed scanner EPSON Perfection® V500 Photo (Epson America Inc., Los Alamitos, CA, USA) and the digitized images were processed with ImageJ software using the Color Histogram plugin [67 (link)]. The obtained results were converted from RGB to CIEL*a*b* values and then the CIEL*a*b* color model was used to evaluate the color of the cookie, since this model has the largest color range and mimics human vision [68 (link)]. The parameter L* represents the luminance or lightness of the sample and ranges from 0 (black) to 100 (white). The chromatic components a* and b* can range from −128 to 127. The parameter a* stands for the green–red and b* for the blue–yellow axis of the color space. The total color difference (∆E) between the control and sample cookies was calculated using the CIE76 color difference equation, which represents the Euclidean distance between two points in CIEL*a*b* space [58 ]. Two sample cookies from every batch were evaluated.
4
Quantifying Brain Infarct Size
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Infarct size was quantified as follows: cryo-embedded brain tissue 5 mm anterior and posterior to the bregma were cut into 10 µm-thick brain slices using a microtome (Leica SM 2000R, Wetzlar, Germany). Every tenth brain slice was stained and digitalized using a scanner (Epson Perfection V500 Photo, Epson, Germany). Brain hemispheres and the area of the lesion were traced manually on each scan using ImageJ Analysis Software v1.52e (National Institutes of Health, Bethesda, MD, USA, https://imagej.nih.gov/ij/ ). The areas were then summed and multiplied by the slice-slice interval.
5
Quantitative Immunoblot and IHC Analysis
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Immunoblots were developed on Fuji RX X-ray film 18 × 24 cm and then scanned on an Epson Perfection V500 Photo flatbed scanner. Images were cropped using Adobe Photoshop, but were otherwise unprocessed. Immunohistochemical staining was imaged on a Zeiss Axio Imager using the Zeiss AxioVision 4.8 software using the AutoLive setting and interactive white balance. Quantification was performed by counting number of cells per field of view for 5 images per organ/mouse, the mean of 5 raw counts was calculated and represents one data point per graph.
6
Lateral Flow Assay Signal Analysis
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Immediately after incubation, RPA reactions were removed from heat and 2 µL was removed and added to 98 µL of tris-buffered saline with 0.05 % tween-20 (TBST), which was used as both sample diluent and lateral flow running buffer. This dilution step is necessary because the dextran sulfate used as a crowding reagent in the RPA nfo buffer causes non-specific binding of the antibody labelled gold to the test line, generating a false positive. After dilution, 10 µL of the product was removed and spotted onto the end of a lateral flow strip. The strip was placed in a well containing 98 µL of TBST and allowed to run for 5 min. The strips were then scanned using a commercial document scanner (Epson Perfection V500 Photo) and images were recorded at 1200 dpi using the ‘reflective document’ settings.
The signal-to-background ratio (SBR) of the lateral flow images were analysed using a custom MATLAB script. The user manually segments the test line and the surrounding background region. The SBR is calculated by calculating the ratio of the average intensity of the signal and background regions [31 (link)].
The signal-to-background ratio (SBR) of the lateral flow images were analysed using a custom MATLAB script. The user manually segments the test line and the surrounding background region. The SBR is calculated by calculating the ratio of the average intensity of the signal and background regions [31 (link)].
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