Flow cytometry determination of VapA cell surface localization in Rhodococcus equi using a C-terminal Strep-tagged VapA.

VapA-ST is expressed as a cell surface protein. Flow cytometry of R. equi ΔvapA carrying either pVapA-ST (A, C and D) or pVapA (B). Approximately 7x10⁵ cells were incubated with THE^2122; NWSHPQFEK Tag mouse FITC-monoclonal antibody (A, B and D). Cells carrying pVapA were used to assess the level of antibody’s non-specific binding (B). The intrinsic fluorescence of R. equi ΔvapA/pVapA-ST was determined by including a control incubated without the antibody (C). The bacterial sample was digested with Trypsin to remove proteins from the cell surface before incubation with antibody (D). Data are a representative of three independent experiments.