Question: When I run the cellranger count pipeline in Antibody-only mode (that is, without gene expression), the .cloupe file is missing from the output folder. I hear that this is related to the number of Cell Surface Proteins (or antibodies/dCODE™ Dextramer®) used in the feature reference file. Could you explain?
Answer: When a cell surface protein library is not accompanied by a gene expression library, Cell Ranger will skip the secondary analysis step if there are less than 10 antibodies/dCODE™ Dextramers® in the feature reference file. As a result, the analysis folder and the .cloupe files are not output. The web summary metrics in the Summary tab and the Feature Barcode matrix are not impacted. Cell Surface Protein data, when run with accompanying gene expression data, is also not affected regardless of the number of features in the feature reference.
10x Genomics does not officially support the analysis of < 10 Cell Surface Proteins without an accompanying gene expression library. If you have less than 10 Cell Surface Proteins, an easy workaround is to pad the feature reference CSV file with additional lines of Cell Surface Proteins (that are not in your experiment). The padded Cell Surface Proteins will return zero UMI counts but enable Cell Ranger to run the complete pipeline, including generating the .cloupe file.
In general, the choice of antibodies in the panel plays a significant role in the effectiveness of cell calling in Feature Barcode only mode. Click here for more details.