Figure 6

Cellular localization of TvBspA625 by indirect immunofluorescence analysis. T. vaginalis cells (G3 isolate) grown in vitro were processed for IFA (ethanol fixation) with the four mouse antisera raised against the four peptides designed from TvBspA625 sequence (see Figure 2) and imaging performed with a confocal microscope. Three antisera (EXT-1, CT-1 and CT-2) gave consistent signals on the cell surface of the parasites whereas EXT-2 often gave strong labelling over most of the cells structures (additional file 18, Figure S7). (A) The mouse antisera raised against a peptide derived from the TvBspA625 cytoplasmic tail (CT-2, green) gave a clear surface labelling. Co-labelling of the hydrogenosomal malic enzyme (red) with a specific rabbit antiserum led to the labelling of intracellular structures as expected [49]. Controls consisted of specific mouse pre-sera or secondary anti-mouse antisera alone (no signal in both cases, data no shown) and a mouse monoclonal antibody raised against tubulin decorated with same secondary anti-mouse antibody (panel C), clearly demonstrated the specificity of the signal attributed to the anti CT-2 peptide antisera. (B) The corresponding DIC image of the panel A. DAPI was added to the mounting media to label the nuclei (blue). The scale bar is shown. (C) Corresponding signal identified for the antisera raised against the TvBspA625 extracellular peptide (EXT-1, green) and processed as in A. (D) The corresponding DIC image of the panel B. (E) Corresponding signal identified with the mouse monoclonal VG2 raised against a T. vaginalis tubulin protein and processed with the same secondary anti-mouse antisera (green) and anti-ME rabbit antisera (red). (F) The corresponding DIC image of the panel E.