10x Genomics Support/Cell Ranger 7.2/Analysis/

Cell Ranger Aggregate Outputs

For convenient multi-sample analysis, the cellranger aggr pipeline generates output files that contain all of the data from the individual input jobs, aggregated into single output files. The GEM well suffix of each barcode is updated to prevent barcode collisions, as described below.

Each output file produced by cellranger aggr follows the format described in the Understanding Outputs section of the documentation, but includes the union of all the relevant barcodes from each input job. For example, samples processed individually using cellranger count and then aggregated using aggr would generate final outputs similar to the count pipeline.

Important
Cell Ranger aggr does not perform a cell calling step, it simply aggregates the cell calls from each input job into a final set of cell calls.

A successful cellranger aggr run should conclude with a message similar to this:

Outputs: - Aggregation metrics summary HTML: /home/jdoe/runs/AGG123/outs/web_summary.html - Copy of the input aggregation CSV: /home/jdoe/runs/AGG123/outs/aggregation.csv - Aggregation metrics summary JSON: /home/jdoe/runs/AGG123/outs/summary.json - Secondary analysis output CSV: /home/jdoe/runs/AGG123/outs/analysis - Filtered feature-barcode matrices MEX: /home/jdoe/runs/AGG123/outs/filtered_feature_bc_matrix - Filtered feature-barcode matrices HDF5: /home/jdoe/runs/AGG123/outs/filtered_feature_bc_matrix.h5 - Loupe Browser file: /home/jdoe/runs/AGG123/outs/cloupe.cloupe Pipestance completed successfully!
Important
Starting with Cell Ranger 6.1, the unfiltered or raw feature-barcode matrix is no longer output by cellranger aggr.

Once cellranger aggr has successfully completed, the outputs can be found in the outs/ sub-directory. The directory structure is shown here:

outs ├── aggregation.csv ├── count │ ├── analysis │ │ ├── clustering │ │ ├── diffexp │ │ ├── pca │ │ ├── tsne │ │ └── umap │ ├── cloupe.cloupe │ ├── filtered_feature_bc_matrix │ │ ├── barcodes.tsv.gz │ │ ├── features.tsv.gz │ │ └── matrix.mtx.gz │ ├── filtered_feature_bc_matrix.h5 │ └── summary.json └── web_summary.html

These files are similar to the cellranger count Gene Expression pipeline outputs. In addition, the aggr pipeline also outputs aggregation.csv, a copy of the input aggregation CSV. The resulting web summary HTML file can be viewed in any supported web browser, and the .cloupe file can be opened in Loupe Browser.

If chemistry batch correction was specified (batch column in input aggregation CSV file), the summary.json and web_summary.html files will contain batch effect scores before and after chemistry correction. Batch effect scores indicate whether there is a batch effect (greater than one) or no batch effect (closer to one) (read more on the gene expression algorithm page).

For more information on summary metrics, refer to the cellranger aggr section of the Summary Metrics page. For aggr datasets with 100k+ cells, Loupe Browser 6.0 and later is recommended for optimal performance.

If your cellranger aggr run includes cell surface protein/Antibody Capture libraries, an additional folder called antibody_analysis is created. The contents of this folder are described on the Antibody Outputs page.

If your cellranger aggr run includes CRISPR Guide Capture libraries, an additional folder called crispr_analysis is created. The contents of this folder are described on the CRISPR outputs page.

Each output file produced by cellranger aggr follows the format described in the Understanding Outputs section of the documentation, and it includes the union of all the relevant barcodes from each input jobs. The GEM well suffix of each barcode is updated to prevent barcode collisions.

Please note that the filtered_contig_annotations.csv includes the donor, origin, and any custom metadata specified in the input aggregation CSV as additional columns.

Here is a list of important files to check in aggr outputs: