PubMed1996
The goal of this research was to synthesize a chimeric ovalbumin/inhibin antigen using recombinant techniques. Antigenic epitopes of the translated product from an ovalbumin cDNA were mapped using computer modeling techniques. The corresponding nucleotide sequences of ovalbumin epitopes were examined for unique restriction sites to allow insertion of a synthetic bovine inhibin alpha gene fragment which encoded the first 26 amino acids. A plasmid (pETOI-8) was synthesized which contained the chimeric ovalbumin/inhibin alpha 1-26 gene. A 46 kDa recombinant protein (ovalin) was produced from BL21 (DE3) Escherichia coli cells containing pETOI-8 that was identified with anti-ovalbumin and anti-inhibin alpha 1-26 antisera. Rabbits were immunized subcutaneously in four sites along the back against ovalbumin (n = 3), crude ovalin (n = 3), or gel-purified ovalin (n = 3) at week 0, 4, 7, and 18. Primary and booster immunizations contained ca 300 micrograms of antigen emulsified in complete Freund's adjuvant and incomplete Freund's adjuvant, respectively. Blood samples collected at week 0, 6, 8, and 19 were evaluated for their ability to bind ovalbumin, 32 kDa bovine inhibin and bovine inhibin alpha 1-26 using ELISA. Mean anti-ovalbumin titers at week 19 were 1:100000 in rabbits immunized against ovalbumin or crude ovalin, and 1:23333 in rabbits immunized against gel-purified ovalin. Anti-inhibin and anti-inhibin alpha 1-26 titers were nonexistent in antisera from pre-immunized rabbits and rabbits immunized against ovalbumin. Mean anti-inhibin titers at week 19 were 1:833 and 1:10000 in rabbits immunized against crude ovalin or gel-purified ovalin, respectively. Mean anti-inhibin alpha 1-26 titers at week 19 were 1:3333 and 1:6666 in rabbits immunized against crude or gel-purified ovalin, respectively. We conclude that genetic engineering of inhibin alpha 1-26 into the antigenic epitopes of ovalbumin provides potential for the development of an anti-inhibin vaccine.