Abstract

The immune system of marine mammals is still largely in its nascent stage of understanding, having been limited to select species for which antibody probes have been developed for research and diagnostics. To date, several antibodies have been identified or developed for immunologic evaluation of manatee tissues. These include anti-manatee immunoglobulin G (IgG) monoclonal (mAB) and polyclonal (pAb) antibodies, an anti-manatee IgM mAb, cross-reactive monoclonals with specificities for a pan T-cell marker (feline) and for macrophages (human Am3-K), and pAB specific for interleukin-2 (human).^1-3 The aim of this study was to identify additional antibodies that could be incorporated into investigations that further define the manatee immune system. Blood samples were collected in ACD vacutainers from free ranging manatees as part of United States Geological Survey-Sirenia Project's annual health assessment studies. These samples were used, via flow cytometry, to identify potential cross-reactivity of 19 monoclonals with specificity for Tursiops, Bovine, Human, Ovine, and Equine cell surface leukocyte differentiation antigens with a focus on lymphocyte markers. While the majority of antibodies tested did not cross react, several exhibited some degree of cross-reactivity. These included antibodies to an uncharacterized pan-leukocyte marker (equine), gamma-delta T cells (bovine) and an MCH class II antigen (bovine). Lymphoid tissue samples (frozen) from the Florida Fish and Wildlife Conservation Commission's Marine Mammal Pathobiology Laboratory are being used to immunohistochemically characterize these cross-reactive antibodies. While the antibodies described above bound native proteins using flow cytometry, none functioned in immunohistology using formalin-fixed tissue sections. Two additional antibodies with specificities for CD3 (pAb; T lymphocytes) and CD20 (B lymphocytes) are currently being tested in formalin-fixed tissues; these latter two antibodies have previously been demonstrated to exhibit broad species specificity in immunohistology using fixed tissues.

Application of the above-mentioned antibodies will begin to allow phenotypic identification of leukocyte subpopulations in manatees. Typically, when species-specific antibodies are not available, antibodies from closely related species can sometimes serve as a substitute. Molecular and skeletal studies have revealed that the manatee is evolutionarily related to the elephant. However, while little is known about the manatee immune system, even less is known about this system in elephants.^4,5 Limited in number, elephant-specific reagents have thus far shown no cross reactivity with manatee tissues. In summary, the antibodies being identified in the current study with cross-reactivity in manatees will undoubtedly further our understanding of this unique and fascinating species. Development of manatee specific antibodies would be most useful for future studies targeting specific leukocyte cell surface markers.

Acknowledgements

The authors would like to thank Dr. Bob Bonde of the United States Geological Survey-Sirenia Project, Pat Lewis of the University of Florida-College of Veterinary Medicine, and Diane Naydan of the University of California Davis-School of Veterinary Medicine. Funding is provided through the Florida Fish and Wildlife Conservation Commission, Whitney Marine Laboratory and the University of Florida Aquatic Animal Health Program, at the University of Florida, College of Veterinary Medicine. This study was conducted under Federal Fish and Wildlife Permit #MA067116-1 and IACUC protocol #200801803 and #200802129.

References

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