Screening and development of monoclonal antibodies for identification of ferret T follicular helper cells

Wenbo Jiang,Daniel S Layton,Stephen J Kent,Adam K Wheatley,Hannah G Kelly,Paul G Whitney,Ian Barr,Jennifer A Juno,Hyon-Xhi Tan,Julius Wong

doi:10.1038/s41598-021-81389-z

Abstract

The ferret is a key animal model for investigating the pathogenicity and transmissibility of important human viruses, and for the pre‐clinical assessment of vaccines. However, relatively little is known about the ferret immune system, due in part to a paucity of ferret‐reactive reagents. In particular, T follicular helper (Tfh) cells are critical in the generation of effective humoral responses in humans, mice and other animal models but to date it has not been possible to identify Tfh in ferrets. Here, we describe the screening and development of ferret-reactive BCL6, CXCR5 and PD-1 monoclonal antibodies. We found two commercial anti-BCL6 antibodies (clone K112-91 and clone IG191E/A8) had cross-reactivity with lymph node cells from influenza-infected ferrets. We next developed two murine monoclonal antibodies against ferret CXCR5 (clone feX5-C05) and PD-1 (clone fePD-CL1) using a single B cell PCR-based method. We were able to clearly identify Tfh cells in lymph nodes from influenza infected ferrets using these antibodies. The development of ferret Tfh marker antibodies and the identification of ferret Tfh cells will assist the evaluation of vaccine-induced Tfh responses in the ferret model and the design of novel vaccines against the infection of influenza and other viruses, including SARS-CoV2.

Highlights

Ferrets (Mustela putorius furo) are a well-established animal model for influenza research and are widely used to investigate the pathogenesis and transmission of influenza viruses and pre‐clinically evaluate the efficacy of influenza v accines[1,2]
Commercial anti-human/mouse BCL6 antibodies were screened for cross-reactivity against ferrets by staining lymph node (LN) cell suspensions recovered from influenza infected ferrets
Identification of T follicular helper (Tfh) cells in ferrets could greatly assist providing an immunological rationale for the design of novel vaccines against the infection of influenza and other viruses, including SARS-CoV2

Summary

Introduction

Ferrets (Mustela putorius furo) are a well-established animal model for influenza research and are widely used to investigate the pathogenesis and transmission of influenza viruses and pre‐clinically evaluate the efficacy of influenza v accines[1,2]. Tfh cells are a subset of CD4 T cells that provide help with B cells for high-affinity antibody production in germinal centres (GC)[12,13,14]. These specialised cells are crucial for the formation of GC, affinity maturation and maintenance of B cell memory. We first identified two commercial anti-human/mouse BCL6 antibodies which had cross-reactivity with ferret lymph node (LN) cells. We developed mouse anti-ferret CXCR5 and PD-1 monoclonal antibodies using single cell PCR-based method. We detected Tfh cells in lymph nodes from influenza infected ferrets using these antibodies

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Results

Conclusion