Export a Graph Pad Prism file directly from OMIQ and create a Grouped plot. The example we give below shows how to quickly go from analysis in OMIQ to two-way ANOVA results in GraphPad Prism.
1. Add Metadata to your Dataset
Add any metadata that is needed to build your Grouped plot. For this example, X-axis (Timepoint) and groups (Sample Type).
You can find out how to add metadata to your datasets here.
2. Build your Workflow
Shown above is just an example (note that the fcs files were already compensated). Build your workflow in whatever way makes sense for the data you wish to export.
In this example exploratory analysis, we have observed that there are less CD4+ T cells in d14 male samples, compared to d14 female samples. Furthermore, there appears to be a difference in the expression level of CCR7 in CD8+ T cells, in d14 male samples, compared to d14 female samples. We will export these parameters, over all timepoints, to see if this is a statistically significant observation.
3. Add a Export Statistics task to your Workflow
Once you have the data needed to build your Grouped plot, click Add new child task and select Export Stats from the task selector.
For demonstration purposes we have added the Export Stats task to the Figure task, though it would be more correct to add this to the upstream Gating task, as the existing branch that has the Figure task has subsampled the data.
You can also access the Export Statistics task from within other tasks, such as the Gating task. Have a look below the Logout button.
4. Setup your Export Statistics task
4.1. Select your Files
Select the Files you want to export statistics for and click Next or click Features.
Most dropdowns in OMIQ allow you to filter the list, just start typing. You can even filter by metadata.
4.2. Select your Features
Select the Filters you want to export Feature Measures for → select the Feature Measures you want to export → select the Features you want the selected Feature Measures of → go to the Counts tab.
In this example, we will export Mean Fluorescent Intensity for CCR7 on our CD4+ and CD8+ cells.
4.3. Select your Counts
Select the Filters you want to export Count Measures for → select the Count Measures you want to export, then skip to Metadata.
In this example, we will export the Percent of Total cells that are CD4+ or CD8+.
4.4. Select your Metadata
It is very important to add the metadata necessary to build your Grouped plot. Select the necessary Metadata Dimensions then go to Configuration.
Go back to Step (1) if you don't see the Metadata you need in the Metadata Dimensions.
4.5. Configure your Export settings
Select Prism in the Export File Format dropdown. Adjust any other settings and click Export or Run Export Stats.
4.6. Download your File
Navigate the the Results tab within the Export Statistics task and click the cloud icon to download your file.
5. Open and Analyze your GraphPad Prism File
5.1. Extract and Rearrange your Multiple Variable Table
Open your GraphPad Prism file and click Analyze → select the Extract and Rearrange analysis and click OK. Change the Format of results table to Grouped for two-way ANOVA → navigate to the Data arrangement tab → define your variables and click OK.
In the example shown, we are viewing the percentage of our total cells that are CD4+ (Response variable), across the sampled Timepoints (Row factor variable), between our Sample Types (Column factor variable). We have then repeated this step for the other response variables of interest (CD8+|percent total, CD4+|BV421-A___CCR7|mean and CD8+|BV421-A___CCR7|mean).
5.2. Perform a Two-way ANOVA Analysis with Multiple Comparisons Test
Completion of the previous step should automatically navigate you to your reshaped table in the Results section. From here, select Two-way ANOVA → select the relevant parameters then click OK. This will generate a new Results table that will open automatically - shown here with the 2way ANOVA results for CD4+|percent_total.
We have then repeated this step for the other tables shown in the Results section.
In this example we have chosen to use the default settings for two-way ANOVA. The best equation will depend on your data and experimental design.
5.3. (Optional) Express your Creative Flair on your Graphs
If the default plot is not to your liking, you can use the many handy tools in GraphPad Prism to adjust it.
In this example, we have adjusted the aesthetics of our Grouped Plots and made a layout. We can see that there were indeed significantly more CD4+ T cells in females, at each timepoint, compared to males. There were no significant differences in any of the other variables tested.
You can share your result easily using Prism Cloud