4 Module list
List and explanation of the different Shiny actions available in the app, along with their functions and usage.
| icon | action | description | prerequite_type | output_type |
|---|---|---|---|---|
|
correct batch effects | 2: fb | 2: fb | |
|
extract features from FCS concat with clustering | 3: sce | 5: fe | |
|
import OMIQ FE | 0: none | 5: fe | |
|
import FCS concat | 0: none | 5: fe | |
|
control quality of features | 5: fe | 6: html | |
|
analyse abundancy of clusters - univar | 5: fe | 6: html | |
|
analyse intensity of clusters - univar | 5: fe | 6: html | |
|
import .rds object | 0: none | 5: fe | |
|
export to phantasus | 5: fe | 4: files |
4.1 Batch Correction
tbd
4.2 Extract Clustering Features
Functionality Overview: This panel allows users to extract features from a specified cell set. Users can define State Markers and choose their preferred clustering method.
Instructions:
-
Specify State Markers:
- Use the provided input field to specify State Markers.
-
Select Clustering Method:
- Choose a clustering method from the dropdown menu. This selection determines how the cells will be grouped.
-
Run Feature Extraction:
- Once you have specified the State Markers and selected a clustering method, click on the "Extract Features" button to initiate the process.
Additional Information:
- Accurate specification of State Markers and the appropriate choice of clustering method are essential for effective feature extraction.
4.3 Import Features
Functionality Overview: The "Import from Feature" panel allows users to import various data tables such as abundance, MFI (Mean Fluorescence Intensity), and metadata tables. Users also have the option to load a marker table. All these tables should be linked by a common ID.
Instructions:
-
Browse and Select Files:
- Click on the ‘Browse’ button to select the data files you wish to import.
- You can import files for abundance data, MFI data, and metadata.
- Optionally, you can also load a marker table. (not available yet)
-
Ensure Common ID Column:
- Make sure that each table has a common ID column. This ID is essential for creating the bound between the different tables.
-
Upload Files:
- After selecting the files, complete widgets.
Additional Information:
- This import functionality is designed especially for OMIQ exported features.
4.4 Import Concatenated FCS
Functionality Overview: This panel allows users to import a concatenated Flow Cytometry Standard (FCS) file representing a complete experiment. Each FCS file within the concatenated file should be annotated and assigned to specific biological samples, studied conditions, etc. Additionally, users must provide a metadata file (in .xlsx or .csv format) containing an FCS ID to identify the origin of cells.
Instructions:
-
Uploading Files:
- If using this application on a server, first upload your files.
- Click on the ‘Browse’ button (max file size: 500MB) to select and upload your concatenated FCS file and metadata file.
-
Selecting FCS Directory:
- After uploading, select the ‘uploaded_fcs’ directory containing the fcs file by clicking on ‘Select your FCS directory’.
-
Metadata File Upload:
-
Upload the metadata file that includes the FCS IDs. This file should be in either
.xlsxor.csvformat. - Ensure that the metadata file correctly maps each FCS ID to its respective sample and condition.
-
Upload the metadata file that includes the FCS IDs. This file should be in either
File Requirements:
- Concatenated FCS file must include properly annotated data with clustering and origine fcs id of each cells.
-
Metadata file in
.xlsxor.csvformat, containing FCS IDs for cell origin identification.
Marker Selection:
- Once the concatenated FCS is loaded, manually select the marker of interest by clicking in the table.
4.5 Quality Control
Functionality Overview: The Quality Control Module provides an interactive interface for users to generate customized quality control reports. Users can set parameters, define thresholds, and choose various options for their report.
Instructions:
-
General Parameters:
- In the ‘General Parameters’ tabset, you can use the widgets to choose general annotations for your report.
-
Threshold Settings:
- Navigate to the ‘Threshold’ tabset to set cutoffs for your analysis, like the minimum number of cells for each cluster.
-
ON/OFF Options:
- In the ‘ON/OFF’ tabset, use boolean widgets to customize your report.
- Options may include showing the code used for analysis or ordering the graphs in a specific manner.
-
Run the Report:
- After setting all parameters and options, click on the "RUN THE REPORT" button located at the top of the page.
- This will initiate the generation of your report.
- The report will take some time to generate so be patient, it will open in a new window.
4.6 Differential Abundance
Functionality Overview: The Differential Analysis Panel is designed for users to conduct comprehensive differential analyses. It includes advanced settings for thresholds, like p-value and fold change, and a dedicated tab for setting univariate parameters to define the differential analysis design.
Instructions:
-
General Parameters:
- Define the Condition column on which the differential comparison is to be performed.
-
Threshold Settings:
- In the ‘Thresholds’ tabset, set critical values for your analysis.
- This includes specifying a p-value threshold for statistical significance and a fold change threshold to identify meaningful differences.
-
Univariate Parameters:
- Navigate to the new ‘Univariate Parameters’ tab to configure the design of your differential analysis. You will need to confirm the condition the ‘Design columns’ selector.
-
Run Analysis:
- After setting all thresholds and univariate parameters, initiate the differential analysis by clicking the "RUN ANALYSIS" button.
4.7 Differential State
tbd
4.8 Import R Object
Functionality Overview: This panel enables users to import an existing .rds file that corresponds to a previously processed Flow Cytometry Standard (FCS) file.
Instructions:
- Browse File: Click on the ‘Browse’ button to navigate through your local directory.
-
Select File: Look for the
.rdsfile you wish to import. These files represent pre-processed FCS data. - Confirmation: Once the file is successfully uploaded, a confirmation message will be displayed.
- Next Steps: After importing, you can proceed to new actions in the "Select an action" box.
Additional Information:
- This panel is designed to streamline the workflow by allowing easy access to pre-processed data.
4.9 Export to Phantasus
Functionality Overview: This panel facilitates the export of your data to Excel format for advanced analysis in Phantasus.
Downloadable Excel Files:
- MFI Split by Markers: Export the MFI data, providing detailed insights split by each marker.
- Median of MFI: Obtain the median MFI values for each FCS file and marker.
- Abundance Matrix: Generate an abundance matrix displaying the percentage of cells in each cluster for each FCS file.