A Customizable Suite of Methods to Sequence and Annotate Cattle Antibodies

Area of Science:

• Immunology
• Genomics
• Veterinary Science

Background:

• Understanding antibody responses is crucial for vaccine and therapeutic development.
• High-throughput sequencing and immunoinformatics tools enable detailed antibody repertoire analysis.
• Previous methods lacked comprehensive high-resolution analysis in cattle.

Purpose of the Study:

• To detail a flexible and customizable suite of methods for antibody repertoire sequencing in cattle.
• To adapt existing methods for the 10x Genomics platform for native heavy-light chain isolation.
• To provide a powerful toolkit for high-resolution cattle antibody response studies.

Main Methods:

• Flow cytometry and single-cell sorting.
• Heavy and light chain amplification and antibody sequencing.
• Adaptation to the 10x Genomics platform and use of the Ig-Sequence Multi-Species Annotation Tool.

Main Results:

• Successfully isolated native heavy-light chain pairs from cattle B cells.
• Sequenced antibody repertoires from three workflows processing 84, 96, and 8313 B cells.
• Obtained 24, 31, and 4756 antibody heavy-light chain pairs, respectively, with discussed throughput, timeline, cost, and equipment considerations.

Conclusions:

• The developed suite of methods provides a powerful toolkit for high-resolution cattle antibody response studies.
• The methods are flexible, customizable, and adaptable to platforms like 10x Genomics.
• The principles can be applied to study antibody responses in other mammalian species.