Figure 1.

Workflow of cattle mAb production using singe B cell antibody techniques. Cattle were sequentially immunized with different FMDV serotype O strains, and then peripheral blood was collected to isolate PBMCs. The isolated PBMCs were stained with a panel of mouse anti-cattle CD21, anti-cattle IgM, and biotinylated antigen, followed by incubation with APC labeled anti-biotin secondary antibody. The single antigen-specific plasmablast was sorted into 96 well-plate with lysis buffer and immediately reverse-transcribed into cDNA. The paired VH and VL sequences was respectively amplified by nested PCR and then sequenced. The variable regions were codon optimized and cloned into pcDNA3.4 expression cassette containing cattle IgG constant region coding sequences. The recombinant mAbs were expressed in CHO-S and purified by affinity and further molecular sieve chromatography.