> For the complete documentation index, see [llms.txt](https://help.connected.illumina.com/llms.txt). Markdown versions of documentation pages are available by appending `.md` to page URLs; this page is available as [Markdown](https://help.connected.illumina.com/dragen-stratamap/dragen-spatial-transcriptome/run-setup/ica-manual-launch.md).
# ICA Manual Launch
### How to Launch a Manual Analysis
1. **Create a Project:** A project can be specific for the DRAGEN Spatial Transcriptome pipeline, or it can contain multiple pipelines and/or tools. For information on creating projects, refer to the Projects section in [Illumina Connected Analytics help](https://help.ica.illumina.com/home/h-projects).
ICA standard storage is used by default as soon as the project is saved. To connect a different storage source, set it up before creating your project. For details and options, refer to the Storage section in [Illumina Connected Analytics help](http://help.ica.illumina.com/home/h-storage).
1. **Upload the sequencing data:** For information on viewing and uploading data, refer to the Data section in [Illumina Connected Analytics help](https://help.ica.illumina.com/project/p-data).
2. **Start Analysis:** In the project, navigate to Pipelines, select the DRAGEN Spatial Transcriptome Pipeline, and then select "Start analysis". Set up the new analysis by configuring the parameters listed in the [table below](#analysis-parameters-on-ica). When the required files are complete, start the analysis.
3. **Review Results:** After analysis is complete, navigate to results in the configured output location.
Please see the Illumina Support Shorts for guidance on how to set up and run DRAGEN Spatial Transcriptome analysis on ICA.
### Analysis Parameters on ICA
To launch an analysis via the ICA user interface, configure a DRAGEN Spatial Transcriptome pipeline analysis with the parameters described below. Parameters are grouped into sections in the ICA UI.
#### Data Inputs
| Parameter Name | Description |
| --------------------------------- | --------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- |
| Run Folder(s) | The run folder(s) containing the files to analyze. BaseSpace defaults to saving sequencing runs into the **ilmn-runs** folder. |
| Samplesheet | Select a sample sheet in CSV format for the analysis. |
| Genome | Select Human, Mouse, or custom genomes in tar.gz format.
See here for more information
|
| Image Files | Select the registered OME-TIFF image files exported from ISIT for analysis. Image files must align with the Samplesheet. |
| Analysis Config Path | (Optional) Select an analysis configuration JSON file to set additional analysis parameters. See the user guide for available options and configuration details. |
| External Segmentation Image Files | (Optional) Use a cell segmentation file generated outside of the pipeline.
See this page for more details.
IMPORTANT:
Files MUST follow the name format
"{SAMPLE\_ID}\_cell\_segmentation\_mask.tif" or "{SAMPLE\_ID}\_nuclei\_segmentation\_mask.tif". Any deviation will cause the workflow to fail.
|
#### General Parameters (all optional)
| Parameter Name | Description |
| --------------------- | ------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- |
| Slide ID | Enter only if not included in the sample sheet. Sample sheets generated using Run Planner already contain the Slide ID. The ID must be found on the slide and match exactly for successful analysis. |
| Run Name | Provide a run name to use in the DRAGEN report and run-level metric files. If not provided, the run name will be the date and time of the run. |
| Compute Resource Size | Dynamic is recommended and will automatically allocate computing resources based on tissue area and flow cell type. Only adjust if troubleshooting — manually select from predefined Medium, Large, or XLarge configurations. Default: `Dynamic`. |
#### Alignment Parameters (all optional)
Parameters controlling read alignment behavior. Additional DRAGEN alignment settings can be configured via the Analysis Config file. See the user guide for available options and configuration details.
| Parameter Name | Description |
| ------------------------------- | ---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- |
| Output BAM | `none` produces no output; `bam` outputs a BAM file. Note: generating BAM alignments will increase compute time and cost. Default: `none`. |
| Include Intronic Reads | By default, reads mapping to intronic regions are filtered out. Enable to include intronic reads in the analysis for capturing unspliced transcripts. See [Exonic and Intronic Alignments](/dragen-stratamap/dragen-spatial-transcriptome/analysis-methods.md#id-1-exonic-and-intronic-alignments) for details. Default: `False`. |
| Use only uniquely aligned reads | When enabled, only reads that uniquely align to the genome are used for analysis; multimapping reads are excluded. By default, multimappers are included by prioritizing alignments to protein-coding genes over non-protein coding genes. See [Multimapping](/dragen-stratamap/dragen-spatial-transcriptome/analysis-methods.md#id-2-multimapping) for details. Default: `False`. |
#### Cell Segmentation Parameters (all optional)
| Parameter Name | Description |
| -------------------------------- | ------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- |
| Output Nuclei Binned Results | Enable to include analysis on nuclei-binned data. Note: full analysis will run on nuclei-binned data, increasing compute time and the size of the output folder. See [Cell and Grid Binning](/dragen-stratamap/dragen-spatial-transcriptome/analysis-methods.md#h-cell-and-grid-binning) for background on binning. Default: `False`. |
| Cell Boundary Expansion Distance | Sets the distance used to expand the nucleus boundary to determine the cell boundary. See [Cell Segmentation](/dragen-stratamap/dragen-spatial-transcriptome/analysis-methods.md#g-cell-segmentation) for details on how expansion works. The default and recommended setting is 5µm. |
#### Tertiary Analysis (all optional)
| Parameter Name | Description |
| -------------- | ------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------ |
| Grid Bin Size | Enter a colon-delimited list of integers to submit additional grid bin sizes in addition to the default of 10µm. Analysis will run on each size, increasing compute time and cost. We do not recommend 3µm or smaller bin sizes. See Grid Binning for details.
Default: 10µm.
|
| Skip Tertiary | Skip PCA, UMAP, and clustering analysis steps to shorten runtime. See [Tertiary Analysis](/dragen-stratamap/dragen-spatial-transcriptome/analysis-methods.md#j-tertiary-analysis) for the steps that are skipped. Default: `False`. |
### ICA-Specific Options
| Parameter Name | Description | |
|---|
| User Reference | The analysis run name. | |
| User Tags | Text labels to help index the analysis. | |
| Notify me when task is completed | (Optional) Receive an email notification when analysis is complete. | |
| Output Folder | The path to the analysis output folder. The default path is the Analyses output folder. | |
| Logs Folder | The path to the analysis log folder. The default path is the Logs output folder. | |
| Subscription | Select the billing account associated with the analysis. | |
| Storage Size | (Optional) Storage Size defaults to the maximum size required to complete analysis. For smaller analyses, a smaller storage size may be selected. 1-2 S4 or 1 25B - Large 3-4 S4 or 2-3 25B - X-Large Anything bigger - 2X-Large | |
For information about using pipelines, refer to [Illumina Connected Analytics support site page](https://support.illumina.com/sequencing/sequencing_software/illumina-connected-analytics.html).
### Imageless Analysis on ICA
For workflows without microscopy images, the DRAGEN Spatial Transcriptome uses the SampleLocator feature to automatically detect sample regions. To run an imageless analysis, use the Imageless DRAGEN Spatial Transcriptome pipeline provided in the DRAGEN Spatial Transcriptome Bundle.
#### How Imageless Mode Works
SampleLocator analyzes sequencing data against the map file to automatically identify and segment tissue regions without requiring microscopy images.
### Data Transfer to Whitelisted Domain and Workgroup
If data is streamed to the incorrect domain (e.g. public or non-whitelisted domain) and is inaccessible in ICA, use the following instructions to transfer data to the correct, whitelisted private domain and workgroup using the BaseSpace Sequence Hub command line interface (BSSH CLI) tool.
These instructions will require you to perform the BSSH CLI tool installation and have your browser open to the intended domain and workgroup while you use a terminal to authenticate the Workgroup.
{% hint style="info" %}
Note: Items that are \[bracketed] require user input.
{% endhint %}
1. Install the BSSH CLI tool onto the device where runs are saved locally according to the instructions found here: [BSSH CLI Installation Instructions](https://developer.basespace.illumina.com/docs/content/documentation/cli/cli-overview)
2. In a browser, enter your private BSSH domain URL and log in.
1. example: https\://\[private domain name].basespace.illumina.com/dashboard
3. Select the whitelisted Workgroup you intend to use and leave your browser open while you move to the next step.
4. Open a terminal and enter the following commands to authenticate your whitelisted Workgroup:
1. Authenticate
`bs auth -c [private domain name] --api-server https://api.basespace.illumina.com`
2. Confirm authentication
`bs -c [private domain name] whoami`
3. If authenticated, the result should look like this with the Workgroup listed under "Name":
INSERT SCREENSHOT HERE
5. Use the following command to send run data to your authenticated Workgroup:
`[location of BSSH CLI] --config [private domain name] run upload --instrument=[NovaSeqX] --name=[run name] --retry [location of run]`
---
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