Immunostaining of Atlantic salmon (Salmo salar L.) leucocytes

Different salmon leucocyte subpopulations were identified by immunostaining using rabbit antiserum raised against the salmonid cell line TO derived from head kidney leucocytes in combination with other available immunoglobulins. The rabbit anti-TO cell line serum immunostained all isolated leucocytes from head kidney, peripheral blood and spleen, as shown by analyses of these leucocytes by flow cytometry and by fluorescence microscopy. In cytospin preparations, the staining of salmon leucocytes using rabbit anti-TO serum as the primary antibody revealed greater morphological details compared to conventional staining procedures, especially among isolated spleen leucocytes where cells with a morphology usually limited to dendritic cells were seen. Other cells of various shapes and protrusions were also stained although the anti-TO serum did not stain protrusions on all cell types. Among the immunoglobulin positive cells, the thin protrusions were only seen when immunostained using anti-IgM antibody. The same was observed for neutrophils stained using the monoclonal E3D9 antibody. The double staining of cells using rabbit anti-TO serum and monoclonal antibodies specific for IgM positive cells or neutrophils clearly show how the morphology of these cells can be compared with the rest of the leucocyte population. The staining of salmon leucocytes by antiserum to a salmon leucocyte cell line TO provides a tool for staining the total population of salmon immune cells, and can be used in immunofluorescence or confocal microscopy in combinations with labelling of cellular components or pathogens. The detailed morphological characteristics, such as cell protrusions, visualized by the presented staining have not been observed on fish leucocytes by conventional cell staining procedures.