Samples sizes used in this study were similar to those routinely used in this field previously. Therefore, no statistical methods were used to additionally predetermine sample sizes. All experiments (cell culture, SCI, and ONC) are routinely performed in our lab. Statistical analyses were conducted using Prism 7 software (GraphPad Software, Inc., La Jolla, CA, USA). Data distribution was assumed to be normal and variances were assumed to be similar, without formal testing. One-sample t-test was used to test single groups. Comparisons of two groups were done by two-tailed unpaired t-test, multiple group comparisons by one-way analysis of variance (ANOVA) with Dunnett’s, Tukey’s, or Sidak’s post hoc test, and two-way repeated measurement ANOVA with Tukey’s or Sidak’s post hoc test. The statistical test and number of in vitro or in vivo experiments used for each analysis are indicated in each figure legend. Data are presented as single data points and means ± SEM. Differences were considered significant when P < 0.05 (*P < 0.05; **P < 0.01; ***P < 0.001; N.S. not significant). Final assembly and preparation of all figures for publication was done using CorelDRAW 2017 (Corel Corporation, Ottawa, Canada).
Coreldraw 2017
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Sourced in Canada, United States
CorelDRAW 2017 is a professional vector graphics software. It provides tools for creating and editing vector-based images, illustrations, and designs.
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Lab products found in correlation
Photo paint 2017, by Corel (4 mentions)
Cell b software, by Olympus (2 mentions)
Focus 6.5.2 pro, by Helicon (2 mentions)
Prism 7, by GraphPad (1 mentions)
Draw graphic suite 2017, by Corel (1 mentions)
Tcs sp8 x, by Leica (1 mentions)
Originpro 9, by OriginLab (1 mentions)
Originpro 2022, by OriginLab (1 mentions)
Sigmaplot, by Grafiti LLC (1 mentions)
Ix83 fluorescence microscope, by Olympus (1 mentions)
Prism 8, by GraphPad (1 mentions)
Cellsens dimension software, by Olympus (1 mentions)
Monarch genomic dna purification kit, by New England Biolabs (1 mentions)
Immunoseq, by Adaptive Biotechnologies (1 mentions)
Immuneaccess database, by Adaptive Biotechnologies (1 mentions)
Paint shop pro 2018, by Corel (1 mentions)
Illustrator cs4, by Adobe (1 mentions)
Stereomicroscope 475 110 9904, by Zeiss (1 mentions)
Phosphoric acid, by Carl Roth (1 mentions)
Tlc scanner, by CAMAG (1 mentions)
16 protocols using coreldraw 2017
1
Detailed Statistical Analysis Protocol
2
Fluorescence Microscopy Analysis of DRG and Spinal Cord
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Dorsal root ganglia and spinal cord sections were visualized using standard fluorescence microscopy (Olympus BX51) and confocal microscopy (Leica TCS SP8X, Wetzlar, Germany). For confocal microscopy, sequential scanning with a white light laser (70% power) tuned to excite AF488 (495 nm laser 47%), AF555 (532 nm 29% intensity), and AF647 (633 nm 53%) was used. Detectors were tuned to emission ranges of 504 to 542 nm for AF488, 550 to 597 nm for AF555, and 649 to 750 nm for AF647. Images (1024 × 1024 pixels) were taken with an oil immersion X20 or X40 objective with a frame average of 6. Spinal cord slices for CTB-labelled projections were optically sectioned (2-µm thick sections) and projected images were reconstructed (50 µm). Images were processed and analyzed using LAS Lite (Leica Microsystems, Wetzlar, Germany), Image J software (NIH), and CorelDraw 2017 (CorelDraw Graphic Suite 2017, Corel) software. Other than making moderate adjustments for contrast and brightness, the images were not manipulated in any way.
3
Statistical Analysis of Biological Data
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For in vitro experiments normalized data were analyzed with a one-sample t-tests, followed by correction for multiple comparisons with the Holm-Bonferroni method, when more than two different conditions were tested relative to the control. Non-normalized data were tested with a paired or an unpaired two-tailed Mann-Whitney test, t-test or a one-way ANOVA test with subsequent Bonferroni correction. in vivo experimental data were analyzed with a Mann-Whitney test. Software used for analyses, processing of data and preparation of graphs were: Excel; GraphPad Prism (version 7 and 8.4.0, GraphPad Software, La Jolla, CA, USA); OriginPro 9.1 (OriginLab Corporation, Northampton, MA, USA); and Corel Draw 2017.(version 19.1.0.419, Corel Corporation, Ottawa, Canada). p < 0.05 was accepted as significant.
4
Biocytin-Filled Neuron Imaging Protocol
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Figures were plotted using OriginPro 2022 (OriginLab). Images of biocytin-filled neurons were adjusted (brightness inversion, contrast, scale bars, z-stack projection) with Fiji software (Schindelin et al., 2012 (link)). Final assembly was achieved using Corel Draw 2017 (Corel Corporation, Ottawa, ON, Canada).
5
Comprehensive Data Analysis Protocol
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Flow cytometry plots were made with FCS Express, while numerical data were compiled in Excel (Microsoft), which was used to calculate means, SEM, and paired t-tests. Bar graphs with SEM were generated in SigmaPlot (Systat Software, v. 6.0) and figures were made in CorelDRAW 2017 (Corel Corporation, v19.1). The raw data supporting the conclusions of this manuscript will be made available by the authors, without undue reservation, to any qualified researcher.
6
Microscopic Analysis of Cell Constructs
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Microscopy of the constructs was performed on days 1, 2, 4, 7 and 14 after printing using an Olympus IX83 fluorescence microscope (Olympus, Tokyo, Japan). Images were taken, merged, and quantified using the Olympus CellSens Dimension software (version 2.2, Olympus, Tokyo, Japan, 2009).
For quantification of cell survival on day 1, we counted the cells in five equal ROIs (regions of interest) in three pictures per sample and calculated the mean cell number per mm2.
Cell proliferation from day 4 to 14 was quantified via the mean gray value intensity of at least three raw images per condition. Respective background fluorescence was deducted by subtraction of the mean background gray value of three ROIs per image, and different exposure times were normalized based on the exposure time of 400 ms with 2fold amplification. The mean gray value intensity of the cells per image was calculated using formula 1:
Protrusions were analyzed with ImageJ by measuring the maximal length of three cells per time point from the center of the cell body to the utmost point of its fluorescence signal.
Graphs were designed using GraphPad Prism 8 (Graph Pad Software, San Diego, CA, USA). Brightness, contrast and intensity of the depicted images were edited with CorelDraw 2017 (Corel Corporation, Ottawa, Canada) for better perceptibility.
For quantification of cell survival on day 1, we counted the cells in five equal ROIs (regions of interest) in three pictures per sample and calculated the mean cell number per mm2.
Cell proliferation from day 4 to 14 was quantified via the mean gray value intensity of at least three raw images per condition. Respective background fluorescence was deducted by subtraction of the mean background gray value of three ROIs per image, and different exposure times were normalized based on the exposure time of 400 ms with 2fold amplification. The mean gray value intensity of the cells per image was calculated using formula 1:
Protrusions were analyzed with ImageJ by measuring the maximal length of three cells per time point from the center of the cell body to the utmost point of its fluorescence signal.
Graphs were designed using GraphPad Prism 8 (Graph Pad Software, San Diego, CA, USA). Brightness, contrast and intensity of the depicted images were edited with CorelDraw 2017 (Corel Corporation, Ottawa, Canada) for better perceptibility.
7
Profiling T-cell Receptor Diversity in Pediatric Epilepsy
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Bulk genomic DNA was isolated from frozen blocks of fresh involved brain tissue from 13 of the 53 pediatric epilepsy surgery cases and whole blood from nine of these cases (Monarch® genomic DNA purification kit, New England Biolabs, Ipswich, MA). Vβ chain TCR sequences were obtained using the ImmunoSEQ® assay (Adaptive Biotechnologies, Seattle, WA). CDR3 sequences were manually curated to remove those that did not start with the third framework cysteine residue at position 104 (14 (link)). Clonality was calculated as described (15 (link)); comparison of proportions Chi-squared tests (16 (link)) were used to determine whether the frequency of the same Vβ CDR3 sequence was significantly different between brain and blood samples from the same patient. Whether Vβ CDR3 sequences corresponded to public T cell clones was determined by searching the immuneACCESS® database (Adaptive Biotechnologies) and similarity to know anti-viral T cell clonotypes was determined using the VDJdb browser (17 (link)). Venn diagrams and heat maps were made with online tools (bioinformatics.psb.ugent.be/webtools/Venn, software.broadinstitute.org/morpheus/). Venn diagrams and heat maps were exported to CorelDraw2017 as scalable vector graphics and portable document format files respectively (Corel Corporation, Ottawa, Canada).
8
External Palp Morphology Imaging Protocol
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All overview images of the external palp morphology were taken using the Visionary Digital Imaging system. All image adjustments were carried out using either Adobe Photoshop CS6 (Adobe systems, Inc., San José, California, USA) or CorelDRAW 2017, Corel PHOTO-PAINT 2017 and Corel PaintShop Pro 2018 (all Corel Corp., Ottawa, Ontario, Canada).
The terminology for the description of the nervous tissue is based on the neuroanatomical glossary by Richter et al. [29 (link)]. Spider-specific terminology is based on the Spider Anatomy Ontology (SPD) [43 ].
The terminology for the description of the nervous tissue is based on the neuroanatomical glossary by Richter et al. [29 (link)]. Spider-specific terminology is based on the Spider Anatomy Ontology (SPD) [43 ].
9
Flax Seedling Stress Response to ABA and Fusarium
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2-Week-old flax seedlings were subjected to 100 μM ABA treatment (sprayed all over the plants) and after 24 h growth they were infected with 1.3 × 10 5 F. oxysporum conidia prepared as described by Di et al. (2017) , and grown for another 4 h. Then the plants were collected and subjected to clearing with 0.15% TCA in ethanol:chloform mixture (4:1, v/v) for 48 h and then submitted to staining with safranin and solophenyl flavine 7GFE (0.1% w/v in 0.1 M Tris/HCl, pH 8.5) as follows. Whole plants were submerged in safranin solution (0.2% w/v safranin in 10% v/v ethanol) for 5 min and then washed three times in water. Subsequently, the plants were stained for 10 min with solophenyl flavine 7GFE (0.1% w/v in 0.1 M Tris/HCl, pH 8.5), and washed again with water (four times) (Knight and Sutherland 2011) . Roots were cutoff and mounted on the slides, observed under the epi-fluorescent microscope Olympus BX50 using UV excitation light (360-370 nm) and documented using the Olympus DP71 camera and Cell^B software (Olympus Optical Co.). Images, taken at different depths, were processed using Helicon Focus 6.5.2 Pro (Helicon Soft Ltd.) and CorelDRAW 2017 (Corel Corporation).
10
Image Enhancing and Composing Protocols
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If necessary, the contrast of images was enhanced using Corel PHOTO-PAINT 2017. The plates were composed with CorelDRAW 2017 (both Corel Corp., Ottawa, Ontario, Canada).
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