Development of a monoclonal antibody against the CD3 epsilon of olive flounder (Paralichthys olivaceus) and its application in evaluating immune response related to CD3 epsilon

Abstract

The T cell receptor (TCR) is the binding site of antigen and is responsible for specifically activating the adaptive immune response. CD3, an essential component of the CD3-TCR complex, is known to be composed of gamma delta and epsilon chains in teleost. However, there are few monoclonal antibodies (mAb) available to identify these molecules on T cells, so we aimed to produce a mAb against CD3 epsilon to improve our understanding of T cell immune response in olive flounder (Paralichthys olivaceus). CD3 epsilon recombinant protein was expressed in yeast, the expression of which was confirmed by SDS-PAGE, MALDI-TOF/TOF MS and Western blot analysis. A CD3 epsilon-specific mAb 4B2 was selected, the specificity of which was examined by confocal microscopy, flow cytometry and RT-PCR, and the mAb was subsequently used to examine the CD3 epsilon lymphocyte population in several different immune organs, with relatively high percentages of these cells seen in trunk-kidney and spleen, while lower percentages were seen in the liver and peripheral blood of olive flounder. During a viral hemorrhagic septicemia virus (VHSV) infection in olive flounder, the number of CD3 epsilon lymphocytes was seen to gradually increase in the liver, spleen and trunk kidney of infected fish until 7 days post infection (dpi). In peripheral blood, on the other hand, the increase in CD3 epsilon lymphocyte numbers peaked by 3 dpi. These results suggest that CD3 epsilon lymphocytes might be involved in the immune response against VHSV. (C) 2017 Elsevier Ltd. All rights reserved.