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When all of the staining was completed for each sample, five sections (centered on the damage area, taking upper left, lower left, upper right, lower right, and intermediate sections) were selected (visualized at 200× magnification) for image acquisition.
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Chromatic aberration in a fluorescence microscope results in slightly different magnifications for images captured at different emission wavelengths [ 38].
Images were captured by laser scanning confocal microscopy at 40X magnification for quantification, images were captured for 10 random fields per coverslip.
The magnification for overviewed images was ×21,000 and the nominal magnification was ×67,000.
For low magnification images, individual images were stitch together using ImageJ.
At least 6 random fields per cross-section were visualized at 20× magnification and used for image analysis that was performed with the NIS-Elements Advanced Research 2.3 imaging software [ 26].
Three fields per tumor at 400× magnification were used for image analysis by Image-Pro Plus software.
This preliminary evaluation of the cell diameters was determined at 40x magnification, using software for image analysis.
The cells positive for cleaved PARP, caspase-3 and cytosolic cytochrome-c were counted using Image Pro-Plus software (v.3.01, Media Cybernetics, Silver Spring, MD, USA) after capturing the image on a Leica microscope (Leica DMLM, Weizlar, Germany) using a 100× magnification for cleaved PARP images and a 40× magnification for caspase-3 and cytochrome-c images.
We have included higher magnification images for the double staining of NeuN and GABAα6 in the insets of the revised Figure 1D.
Higher magnification images for the double staining of BLBP/βgal and NeuN/βgal have also been included as insets in the revised Figure 3B (which corresponds to the old Figure 3D).
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