ABSTRACT
An ELISA for detection of monoclonal antibodies to surface antigens on canine lymphocytes was evaluated, and results were compared with three additional tests: a radioimmunoassay using 125I-labeled staphylococcal protein A (IPA), the fluorescence activated cell sorter (FACS II), and CdL. Three potentially interesting hybridomas were subsequently selected from two fusions.
Subject(s)
Antibodies, Monoclonal/analysis , Antigens, Surface/immunology , Enzyme-Linked Immunosorbent Assay , Immunoenzyme Techniques , Monocytes/immunology , Animals , Antigens, Neoplasm/immunology , Cell Separation , Cytotoxicity Tests, Immunologic , Dogs , Flow Cytometry , Histocompatibility Antigens/immunology , Hybridomas/immunology , Leukemia/immunology , Radioimmunoassay , Staphylococcal Protein A/immunologyABSTRACT
Four murine monoclonal antibodies were used in cytolytic assays to identify cell populations involved in canine T cell effector functions. Antibody DT-2, directed at T cells, and antibody DLy-6, a panlymphocyte antibody, inhibited mixed lymphocyte culture (MLC) responses and cytotoxic lymphocyte (CTL) activity only when lymphocytes were treated before culture (day 0), but they had no significant effect on these functions when cells were treated after 6 days in culture. Antibody DLy-1, reacting with canine lymphocytes and monocytes, abrogated MLC responses and CTL activity when responder cells were treated on day 0. When cells were treated after 6 days in culture, MLC responses were reduced to 47% of control, whereas CTL activity increased slightly. In contrast, the anti-Ia antibody 7.2 significantly reduced MLC responses and CTL activity of cells treated on either day 0 or day 6 of culture, suggesting that canine CTL express Ia antigens.
Subject(s)
Dogs/immunology , T-Lymphocytes, Cytotoxic/immunology , Animals , Antibodies, Monoclonal , Antigens, Surface/analysis , Cell Differentiation , Cytotoxicity, Immunologic , Histocompatibility Antigens Class II/analysis , Lymphocyte Activation , Lymphocyte Culture Test, Mixed , T-Lymphocytes, Cytotoxic/classificationABSTRACT
Canine blood lymphocytes were nonlytically separated on antibody-coated petri dishes into surface immunoglobulin-positive (SIg+) and -negative (SIg-) populations. SIg- cells were further separated into cells reactive or non-reactive with monoclonal antibody DT-2 recognizing canine T lymphocytes. The purity of the three enriched lymphocyte populations exceeded 90% as assessed by immunofluorescence. Mitogen stimulation showed a vigorous response of SIg+ cells to pokeweed mitogen and concanavalin A but only a weak response to phytohemagglutinin. In mixed DT-2- and DT-2+ cells responded to phytohemagglutinin, concanavalin A and pokeweed mitogen, and both populations were good responders in mixed leukocyte culture. Only DT-2- cells were potent stimulators; DT-2+ cells were not. Hence, canine blood T cells can be divided into two subsets, DT-2+ and DT-2-, both of which are responsive to mitogens and alloantigens.
Subject(s)
Antibodies, Monoclonal , B-Lymphocytes/immunology , T-Lymphocytes/immunology , Animals , Cell Separation , Concanavalin A , Dogs , Lymphocyte Activation , Phytohemagglutinins , Pokeweed MitogensABSTRACT
Two murine monoclonal antibodies directed against canine lymphocytes are described. DLy-1, raised against puppy thymocytes, and DLy-6, raised against bronchoalveolar lymphocytes, both react with most lymphocytes in peripheral blood, thoracic duct lymph, and bronchoalveolar lavage fluids. DLy-1 also recognizes monocytes and granulocytes. However, it is not reactive with erythrocytes, megakaryocytes, or platelets. Expression of DLy-1 antigen on thymocytes ranged from 5--30%. The distribution of DLy-6 antigens seems to be confined to lymphoid cells. Ten to 60% puppy thymocytes were positive. Interestingly, lymphoblasts formed in response to stimulation with mitogens or alloantigens lacked DLy-6 in contrast to DLy-1 cell surface antigen expression.
