The IC50 values were determined by using a linear regression procedure (OriginPro 2018b software, OriginLab Corporation, Northampton, MA, USA). PCA was employed on spectral data as an unsupervised multivariate approach (OriginPro 2018b software); all the PCA comparisons were performed separately on the selected regions of interest (ROI) 3050–2800 cm−1 and 1300–900 cm−1. Normally distributed data derived from FTIRM spectra were presented as mean ± S.D; significant differences between experimental groups were determined by means of a factorial analysis of variance (one-way ANOVA), followed by Tukey’s multiple comparisons test, by the statistical software Prism6 (Graphpad Software, Inc., San Diego, CA, USA); One-way ANOVA compares the means of OSCC-Ctrl-24, OSCC-Ctrl-48, OSCC-5Aza-24, OSCC-5Aza-48, CSC-Ctrl-24, CSC-Ctrl-48, CSC-5Aza-24, and CSC-5Aza-48 groups in order to make inferences about the population means; statistical significance was set at p < 0.05; different letters over histograms indicate statistically significant differences among the above defined experimental groups.
Originpro 2018b software
Manufactured by OriginLab
Sourced in United States
OriginPro 2018b is a data analysis and graphing software developed by OriginLab. It provides tools for data processing, visualization, and statistical analysis. The software is designed for scientific and engineering applications, offering a wide range of features to handle various data types and create publication-quality graphs.
Automatically generated - may contain errors
Lab products found in correlation
Prism v8, by GraphPad (2 mentions)
Prism 6, by GraphPad (1 mentions)
Nano s, by Malvern Panalytical (1 mentions)
Invenio ftir spectrometer, by Bruker (1 mentions)
Opus 7.5 software package, by Bruker (1 mentions)
Platinum atr, by Bruker (1 mentions)
Opus 7, by Bruker (1 mentions)
4 to 20 mini proteantgx precast gels, by Bio-Rad (1 mentions)
Silverxpress silver staining kit, by Thermo Fisher Scientific (1 mentions)
β mercaptoethanol, by Bio-Rad (1 mentions)
Laemmli sample buffer, by Bio-Rad (1 mentions)
Originpro 2018b, by OriginLab (1 mentions)
10 protocols using originpro 2018b software
1
FTIRM Analysis of OSCC and CSC
2
Statistical Analysis of Spectral Data
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Statistical analyses were performed using Graph Pad Prism V8.0.1. (GraphPad Software, Inc., San Diego, CA, USA). The normality of the data was checked using the Shapiro–Wilk test and the appropriate statistical assay was accordingly applied. For parametric data an unpaired t test with Welch’s correction was used, whereas, for non-parametric data, a Mann–Whitney test was applied. Statistical significance was set at p < 0.05 for all the tests.
Regarding the IR data, the differences between groups were statistically analyzed with the software package Graph Pad Prism V8.0.1. (GraphPad Software, San Diego, CA, USA). All data were presented as mean ± SD. Statistical significance among groups was evaluated using Student’s t-test. Statistical significance was set at p < 0.05. Moreover, a PCA was performed as an unsupervised multivariate approach to compare the spectral profiles of CTRL and PRE experimental groups (OriginPro 2018b software, OriginLab Corporation, Northampton, MA, USA) [29 (link),31 (link)].
Regarding the IR data, the differences between groups were statistically analyzed with the software package Graph Pad Prism V8.0.1. (GraphPad Software, San Diego, CA, USA). All data were presented as mean ± SD. Statistical significance among groups was evaluated using Student’s t-test. Statistical significance was set at p < 0.05. Moreover, a PCA was performed as an unsupervised multivariate approach to compare the spectral profiles of CTRL and PRE experimental groups (OriginPro 2018b software, OriginLab Corporation, Northampton, MA, USA) [29 (link),31 (link)].
3
Statistical Analysis of Experimental Data
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All data analysis and plots were done by OriginPro 2018b software (Northampton, MA, USA). All data were subjected to Shapiro-Wilk normality testing, normally distributed data were analyzed by one way ANOVA and non-normal distributed data were analyzed by non-parametric Mann-Whitney test. Results are expressed as mean ± SEM and the threshold for statistical significance (p-value) was set at 0.05. All the experiments were repeated at least three times.
4
Seasonal and Leaf Removal Impacts on Plant Compounds
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The OriginPro 2018b software (OriginLab, Northampton, MA, USA) was used to perform the ANOVA and MANOVA analyses to assess the impacts of the season (S), leaf removal (LR) and their interaction. The S and LR interactions of the principal and significant chemical, phenol, anthocyanin and volatile compounds were reported and discussed. All analyses were repeated three times for each sample.
5
Curve Fitting and Statistical Analysis
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Standard curve and equation derivation were determined from logarithmic curve fitting in OriginPro 2018b Software (OriginLab, Northampton, MA, USA). Differences across three timepoints were determined using 1-way ANOVA. Differences between two categories were determined using the non-parametric Mann–Whitney test. Relationship between two continuous variables were determined using the non-parametric Spearman’s correlation. P values < 0.05 were considered significant. All tests were performed using GraphPad Prism v8.0 (San Diego, CA, USA).
6
Statistical Analysis of Spectral Profiles
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Graph Pad Prism V8.0.1. (GraphPad Software, Inc., San Diego, CA, USA) was used to perform the statistical analyses. Once the normality of the data was assessed by the Shapiro–Wilk test, the following statistical analyses were performed: unpaired t-test with Welch’s correction for the parametric data and Mann–Whitney for non-parametric data. In all cases, the statistical significance was set at p < 0.05.
Concerning multivariate analysis of IR data, the processed spectra were converted in second derivative mode (Savitzky–Golay filter, 9 points of smoothing) and subjected to multivariate analysis, without any other further pre-processing. An unsupervised multivariate approach, Principal Components Analysis (PCA), was used to compare the spectral profiles of CTRL and GDM, and CTRL and SGA experimental groups (OriginPro 2018b software, OriginLab Corporation, Northampton, MA, USA) [23 (link),25 (link)].
Concerning multivariate analysis of IR data, the processed spectra were converted in second derivative mode (Savitzky–Golay filter, 9 points of smoothing) and subjected to multivariate analysis, without any other further pre-processing. An unsupervised multivariate approach, Principal Components Analysis (PCA), was used to compare the spectral profiles of CTRL and GDM, and CTRL and SGA experimental groups (OriginPro 2018b software, OriginLab Corporation, Northampton, MA, USA) [23 (link),25 (link)].
7
Zeta Potential and Carboxyl Content Analysis
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Zeta potential (Φz) was measured in samples with solid concentration of 0.05% w/w (pH = 6.0, 25 °C) using a Zeta potential equipment (Malvern Panalytical, Zetasizer Nano S, Worcestershire, UK), and charge (ζ) measurements were carried out by conductometric titration.
Carboxyl content of the cellulosic samples was analyzed by conductometric titration according to the standard SCAN-CM 65:02 [49 ]. The titration was executed using an automatic titration device (Methrom 751 GPD Titrino and Tiamo 1.2.1 software, Metrohm AG., Herisau, Switzerland). The data of the titration was processed with OriginPro 2018b software (OriginLab Corporation, Northampton, MA, USA). A blank sample (water) was used to exclude systematic error within the analysis.
Carboxyl content of the cellulosic samples was analyzed by conductometric titration according to the standard SCAN-CM 65:02 [49 ]. The titration was executed using an automatic titration device (Methrom 751 GPD Titrino and Tiamo 1.2.1 software, Metrohm AG., Herisau, Switzerland). The data of the titration was processed with OriginPro 2018b software (OriginLab Corporation, Northampton, MA, USA). A blank sample (water) was used to exclude systematic error within the analysis.
8
Infrared Analysis of Dental Aligners
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The infrared analysis of the aligners was carried out with a Bruker Invenio FTIR spectrometer equipped with a Platinum ATR accessory for reflectance measurements (OPUS 7.5 software package, Bruker Optics, Ettlingen, Germany).
The ATR-FTIR spectra were collected at room temperature on dried samples just before the in vitro aging treatments (considered as a control group, Ctrl) and at the time points of 24 and 48 h. Five spectra were acquired in reflection mode in the spectral range 4000–600 cm−1 (spectral resolution 4 cm−1, 64 scans) on the external surface of each sample. A background spectrum was collected on the clean diamond crystal, before each sample measurement. All IR spectra were converted into Absorbance mode, interpolated in the spectral range 1800–700 cm−1, and two-points baseline linear fitted (OPUS 7.5 software, Bruker Optics). These preprocessed spectra were then subjected to principal component analysis (PCA; OriginPro 2018b software; OriginLab Corporation, Northampton, MA, USA) [25 (link)].
For each experimental group, height values of specific bands were calculated by using the Integration routine (K mode) (OPUS 7.5 software package, Bruker Optics, Ettlingen, Germany).
The ATR-FTIR spectra were collected at room temperature on dried samples just before the in vitro aging treatments (considered as a control group, Ctrl) and at the time points of 24 and 48 h. Five spectra were acquired in reflection mode in the spectral range 4000–600 cm−1 (spectral resolution 4 cm−1, 64 scans) on the external surface of each sample. A background spectrum was collected on the clean diamond crystal, before each sample measurement. All IR spectra were converted into Absorbance mode, interpolated in the spectral range 1800–700 cm−1, and two-points baseline linear fitted (OPUS 7.5 software, Bruker Optics). These preprocessed spectra were then subjected to principal component analysis (PCA; OriginPro 2018b software; OriginLab Corporation, Northampton, MA, USA) [25 (link)].
For each experimental group, height values of specific bands were calculated by using the Integration routine (K mode) (OPUS 7.5 software package, Bruker Optics, Ettlingen, Germany).
9
SDS-PAGE Analysis of Silk Nanoparticles
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The silk protein size
distribution was assessed using sodium dodecyl sulfate polyacrylamide
gel electrophoresis (SDS-PAGE). Lyophilized silk nanoparticles were
dissolved in 9.3 M LiBr solution at 60 °C and dialyzed (molecular
weight cutoff 3500 Da) against ultrapure water for 24 h to remove
the LiBr salt. Laemmli sample buffer and β-mercaptoethanol (Bio-Rad
Laboratories, Inc., Hercules, CA, U.S.A.) were added, and the samples
were reduced for 10 min at 70 °C and loaded onto 4 to 20% Mini-PROTEAN
TGX precast gels (Bio-Rad Laboratories). The gels were run with trisglycine
SDS running buffer under reducing conditions. Spectra Multicolor High
Range Protein Ladder (Invitrogen, Carlsbad, CA, U.S.A.) was run as
a molecular standard. The gels were stained with a SilverXpress Silver
Staining Kit (Thermo Fisher Scientific, Waltham, MA, USA). Images
were acquired with an 8 megapixel Wileyfox Swift 2 plus phone, and
the pixel density was analyzed by ImageJ v1.52n (National Institutes
of Health, Bethesda, MD, U.S.A). The vertical profile was plotted
against the distance in pixels using OriginPro2018b software (OriginLab
Corporation, Northampton, MA, U.S.A.).
distribution was assessed using sodium dodecyl sulfate polyacrylamide
gel electrophoresis (SDS-PAGE). Lyophilized silk nanoparticles were
dissolved in 9.3 M LiBr solution at 60 °C and dialyzed (molecular
weight cutoff 3500 Da) against ultrapure water for 24 h to remove
the LiBr salt. Laemmli sample buffer and β-mercaptoethanol (Bio-Rad
Laboratories, Inc., Hercules, CA, U.S.A.) were added, and the samples
were reduced for 10 min at 70 °C and loaded onto 4 to 20% Mini-PROTEAN
TGX precast gels (Bio-Rad Laboratories). The gels were run with trisglycine
SDS running buffer under reducing conditions. Spectra Multicolor High
Range Protein Ladder (Invitrogen, Carlsbad, CA, U.S.A.) was run as
a molecular standard. The gels were stained with a SilverXpress Silver
Staining Kit (Thermo Fisher Scientific, Waltham, MA, USA). Images
were acquired with an 8 megapixel Wileyfox Swift 2 plus phone, and
the pixel density was analyzed by ImageJ v1.52n (National Institutes
of Health, Bethesda, MD, U.S.A). The vertical profile was plotted
against the distance in pixels using OriginPro2018b software (OriginLab
Corporation, Northampton, MA, U.S.A.).
10
Identifying Inhibitory Effects in Anaerobic Digestion
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One-way analysis of variance (ANOVA) followed by Tukey Post-Hoc Analysis (p < 0.05) was carried out to determine the statistically significant differences among mono-and co-digestion samples using the OriginPro 2018b software (OriginLab Corporation, USA). IC50 of potassium inhibition was calculated by nonlinear regression analysis (DoseResp Model) using the OriginPro 2018b software (OriginLab Corporation, USA). Synergistic and antagonistic effects of co-digestion was calculated according to (Tsapekos et al., 2018) . The coefficient of determination (R 2 ) and root mean square error (RMSE) were calculated by the OriginPro 2018b software (OriginLab Corporation, USA).
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