A recombinant form of the outer membrane protein (A-layer protein) associated with atypical Aeromonas salmonicida was expressed, fused to a cellulose binding domain (CBD) isolated from Clostridium cellulovorans. The resultant chimerical protein was bound to either Sigmacell 20® or Orbicell™ cellulose particles. Common goldfish were injected intraperitoneally with the cellulose-protein complex and blood serum antibody levels produced against A-protein were examined weekly by means of ELISA. These titers were compared to those induced by immunization of goldfish with the same protein, with or without Freund's incomplete adjuvant, as well as to a standard bacterin-adjuvant system. Small Orbicell beads (1-10μM) induced antibody levels that were equal to the titers produced by the adjuvanted protein and bacterin formulae. In comparison, the larger Sigmacell particles (10-20μM) proved to be poor immunopotentiators. The long-term titer elicited from a single injection of A-protein bound to Orbicell beads was equivalent to that induced by two injections. All the vaccinated fish demonstrated memory to the A-layer protein after exposure to a pathogenic load of atypical A. salmonicida with Orbicell treated fish displaying the highest titer. No direct correlation was found between the presence of anti-A-protein antibodies and protection against infection. The paper describes a simple and safe method to increase the potential immunogenicity of soluble recombinant proteins by employing relatively inexpensive cellulose particles.
Bibliographical noteFunding Information:
This work was supported by grants from the Israeli Ministry of Industry and Commerce and The Hebrew University of Jerusalem (Agreement no. 0325878). The authors would like to thank David Shelach for expert maintenance of the wet-lab and the experimental fish and to Paulina Mendoza and Dr. Noah Lavid for the weekly collection of blood samples.
- Cellulose binding domain (CBD)
- Recombinant A-protein