Immunolocalization of Fluke Protein FhKT1.1

Adult flukes were fixed in paraformaldehyde at 4 °C overnight, washed with PBS, dehydrated with ethanol and embedded in JB-4 resin (Sigma-Aldrich)^{4 (link),10 (link)}. Sections (2 μm) were incubated in either an anti-peptide antibody raised in mice to the following FhKT1.1 amino acid sequence, Cys-Glu-Gly-Asn-Asp-Asn-Arg-Phe-Asp-Ser-Lys-Ser-Ser-Cys, or pre-immune sera, each at a 1:500 dilution. Sections were then washed three times in PBS with 0.5% Triton X-100 for 30 min and incubated in a 1:500 dilution of the secondary antibody, fluorescein isothiocyanate (FITC)-labelled goat anti-mouse immunoglobulin (Sigma-Aldrich). After three washes in PBS with 0.5% Triton X-100 for 30 min sections were dried and coverslips mounted using glycerol:PBS (9:1) containing 100 mM propyl gallate. Sections were viewed using a Leica DM 2500 light microscope under the HCX PL FLUOSTAR × 10 and × 40 lenses.