Optimized Multicolour Immunofluorescence Panel for Cattle B Cell Phenotyping by an 8-Colour, 10-Parameter Panel

Abstract

AbstractThis 8-colour, 10-parameter panel has been optimised to distinguish between functionally distinct subsets of cattle B cells in both fresh and cryopreserved peripheral blood mononuclear cells (PBMCs). Existing characterised antibodies against cell surface molecules (immunoglobulin light chain (S-Ig(L)), CD20, CD21, CD40, CD71 and CD138) enabled the discrimination of 24 unique populations within the B cell population. This allows the identification of five putative functionally distinct B cell subsets critical to infection and vaccination responses; 1) naïve B cells (BNaïve), 2) regulatory B cells (BReg), 3) memory B cells (BMem), 4) plasmablasts (PB) and 5) plasma cells (PC). Although CD3 and CD8α can be included as an additional dump channel, it does not significantly improve the panel’s ability to separate “Classical” B cells. This panel will promote better characterisation and tracking of B cell responses in cattle as well as other bovid species as the reagents are likely to cross react.