The Single Cell Antibody Annotator operation is an alternate way to annotate and analyze the variable regions of standard IgG-like molecules. It combines the sequences analyzed into fewer representative sequences and can pull out the dominant chain(s) within your dataset.
The Single Cell Analysis operation is particularly suited for analysing Barcoded Sequences from both "single clones in wells" and "single cells in droplets" experiments. It is also suitable for analyzing datasets from NGS technologies that incorporate UMIs. To learn more about Barcodes and UMIs, see this article: Understanding Single Cell technologies: Barcodes and UMIs. The video below provides an overview of these concepts:
For Barcode and/or UMI analysis, you will first need to run the Collapse UMI Duplicates and Separate Barcodes tool. For more discussion about possible workflows, see the Single Cell Analysis workflows article.
- How do I run Single Cell Antibody Annotator?
- Single Cell Antibody Annotator Options
How do I run Single Cell Antibody Annotator?
Select one or more nucleotide sequences or lists in your folder and select Annotation > Single Cell Antibody Anotator:
To run the Single Cell Antibody Annotator operation, select options relevant to your sequence data in the dialog that appears and click Run.
Each selected input sequence list will be analysed independently. The pipeline works by:
- Trimming sequences and then applying filtering steps to reduce total data size (configurable)
- De-novo assembling reads together (optional)
- Identifying and annotating Ig-like Antibody regions (FR/CDR)
- Collapsing sequences with similar V(D)J regions into a single dominant sequence. Heavy and light chains are compared separately.
When run on barcoded sequences in combination with the Collapse UMI Duplicates and Separate Barcodes operation, it can identify heavy-light chains pairs and determine dominant chains within each cell/barcode. For an example of a workflow for analysing barcoded data, please refer to the UMI/Barcode and Single Cell Analysis tutorials.