Materials and Methods
Samples of the F and RBC2 lines were compared for their responses to two mitogens, concanavalin A (Con A) and phytohemagglutinin (PHA)-M. The mitogenic responses were examined for purified peripheral blood mononuclear cells and whole blood. For Con A and PHA-M, each bird was injected with 25 µg/mL and 100 µg/mL, respectively, of the mitogen. Blood samples were collected prior to injection and at 2, 4, 7, and 14 days post-injection.
To measure turkey CD4 and CD8 T cell levels, the cross-reactivity of mouse anti-chicken CD4 and CD8 monoclonal antibodies (mAb) with turkey leucoytes was tested by flow cytometric analysis of blood obtained from individuals in five turkey lines (F, RBC2, and three commercial sire lines). Peripheral blood lymphocytes were isolated and stained with single or dual color staining.
Serum IgG and IgM concentrations in serum were measured in 6-week-old individuals from the F and RBC2 lines prior to challenge with P. multocida. Concentrations of the immunoglobulins were quantitatively estimated with a Sandwich ELISA technique using the cross-reactive polyclonal antibodies specific for the chicken Ig isotypes.
Other evaluations of the immune system of the F and RBC2 lines included: 1) the primary and secondary antibody responses to several antigens (sheep red blood cells, Brucella abortus, P. multocida, and Newcastle disease virus); 2) weight of lymphoid organs (spleen and bursa of Fabricius; and 3) phagocytic activity. Antibody responses to the various antigens were evaluated by a protocol similar to that reported by Li et al. (2000b). Phagocytic activity was measured by carbon clearance from the blood. The supernatant fraction of India ink was obtained through centrifugation and injected into the brachial vein at a volume of 1.5 mL/kg of body weight. Blood was taken from the opposite wing before injection and at 3, 6, and 15 minutes after the carbon injection. After brief centrifugation, the relative amount of carbon particles remaining in the supernatant was spectrophotometrically measured.