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A Customizable Suite of Methods to Sequence and Annotate Cattle Antibodies

by Nyagwange, James , Warimwe, George M. , Charleston, Bryan , Freimanis, Graham , Nzau, Benjamin , Hammond, John A. , Bonnet-Di Placido, Marie , Ramirez Valdez, Kristel , Schwartz, John C. , Jarman, Michael , Morrison, Liam J. , Steyn, Angela W. , Mwangi, William , Dorey-Robinson, Daniel

in 10x Genomics / Amino acids / Annotations / Antibodies / antibody discovery / antibody repertoire / Antibody response / antibody sequencing / Antigens / Cattle / Cloning / Disease / Equipment costs / Flow cytometry / Gene amplification / Genetic aspects / High resolution / High-throughput screening (Biochemical assaying) / Identification and classification / IgMAT / Immune response / Immunoglobulins / Immunological research / Invoices / Light / Lymphocytes B / Methods / Pathogens / Physiological aspects / Polymerase chain reaction / Viral antibodies

2023

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A Customizable Suite of Methods to Sequence and Annotate Cattle Antibodies

2023

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2023

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Journal Article

A Customizable Suite of Methods to Sequence and Annotate Cattle Antibodies

Nyagwange, James,

Warimwe, George M.,

Charleston, Bryan,

Freimanis, Graham,

Nzau, Benjamin,

Hammond, John A.,

Bonnet-Di Placido, Marie,

Ramirez Valdez, Kristel,

Schwartz, John C.,

Jarman, Michael,

Morrison, Liam J.,

Steyn, Angela W.,

Mwangi, William,

Dorey-Robinson, Daniel

2023

Overview

Studying the antibody response to infection or vaccination is essential for developing more effective vaccines and therapeutics. Advances in high-throughput antibody sequencing technologies and immunoinformatic tools now allow the fast and comprehensive analysis of antibody repertoires at high resolution in any species. Here, we detail a flexible and customizable suite of methods from flow cytometry, single cell sorting, heavy and light chain amplification to antibody sequencing in cattle. These methods were used successfully, including adaptation to the 10x Genomics platform, to isolate native heavy–light chain pairs. When combined with the Ig-Sequence Multi-Species Annotation Tool, this suite represents a powerful toolkit for studying the cattle antibody response with high resolution and precision. Using three workflows, we processed 84, 96, and 8313 cattle B cells from which we sequenced 24, 31, and 4756 antibody heavy–light chain pairs, respectively. Each method has strengths and limitations in terms of the throughput, timeline, specialist equipment, and cost that are each discussed. Moreover, the principles outlined here can be applied to study antibody responses in other mammalian species.

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Publisher

MDPI AG,MDPI

Subject

/ antibody repertoire

/ Antibody response