Title

An Analysis of the Humoral Immune System of Three Species of Cetaceans: The Atlantic Bottlenose Dolphin (Tursiops truncatus), Beluga Whale (Delphinapterus leucas), and Pacific White-Sided Dolphin (Lagenorhynchus obliquidens)

Date of Award

1998

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Biological Sciences

First Advisor

Bobby L. Middlebrooks

Advisor Department

Biological Sciences

Abstract

Very little information is available concerning the immune systems, especially humoral immune system, of cetaceans. In a preliminary study of how a cetacean's humoral immune system responds to an immunogen, four Tursiops truncatus (Atlantic bottlenose dolphin) were employed in two immunization protocols using bovine serum albumin (BSA) as the immunogen. The primary and secondary immune responses in these animals were monitored by a double-indirect enzyme linked immunosorbent assay (ELISA) to measure the antibodies produced against the BSA. The results from the ELISA clearly showed that the immunization protocol with multiple injections was more effective at producing an immune response in these animals than the one using a single injection. However, no antibodies were detectable after six months with either of the immunization protocols. Qualitative characterization of the primary and secondary immune response was not possible because of the limitations of the anti-T. Truncatus antibodies used in the ELISA; for such characterization it was necessary to produce antibodies specific for individual immunoglobulin classes of each cetacean species to be studied. In order to provide these antibodies, the focus of the study shifted to the development of immunochemical techniques to isolate and purify the three major immunoglobulin isotypes involved in humoral immunity (IgG, IgA, and IgM) from the serum of T. truncatus, Delphinapterus leucas (beluga whale), and Lagenorhynchus obliquidens (Pacific white-sided dolphin). It was determined that IgG could be purified using a Protein G Sepharose$\sp{\circler}$ 4 Fast Flow affinity chromatography column, that IgA could be purified using an Immobilized Jacalin affinity chromatography column, and that IgM could be purified using a Sephacryl$\sp{\circler}$ S400-HR gel filtration chromatography column. IgM has only been purified from T. truncatus thus far due to limited sources of sera for the other two cetacean species. These purified immunoglobulin isotypes were partially characterized, and antisera were produced against several of the isotypes. To date antisera have been produced against (1) T. truncatus whole serum, precipitated serum, whole IgG, and gamma ($\gamma$) heavy chains; (2) D. leucas precipitated serum and $\gamma$ heavy chains; and (3) L. obliquidens precipitated serum and $\gamma$ heavy chains.