Preparation and Application of Rabbit Polyclonal Antibodies against Elizabethkingia meningosepticum on Frogs

  Objective  Rabbit polyclonal antibodies against Elizabethkingia meningosepticum (Em) were prepared to establish an indirect ELISA detection to trace the pathogenic attack in organs of frogs (Rana spp).

  Method  The strain RsB1151018NA of Em from a diseased R. spinosa was isolated, formalin-inactivated, and injected into a rabbit for the serum preparation. An optimized indirect ELISA using the rabbit polyclonal antibodies was developed for tracing RsB1151018NA. The rabbit immunoglobulin G was purified on a protein A column. The purified antibodies were marked with FITC and further purified on a desalting column Sephadex G-25 for subsequently tracing the pathogen in organs of the inoculated frogs.

  Results  The obtained rabbit polyclonal antibodies showed a titer up to 1 ∶ 5.12×10⁵, and 1 ∶ 1.6×10⁴ after purification. The indirect ELISA assay on the polyclonal antibody was specific and sensitive in detecting Em without cross-reaction on other bacteria. It had a detection limit of 1.0×10⁴ cfu·mL⁻¹. According to the in vivo tracing test, RsB1151018NA distributed throughout the entire frog body by blood circulation with the most serious infections in the kidney, eyes, and brain.

  Conclusion  The optimized indirect ELISA detection of Em was rapid, sensitive, and specific in tracing the bacterial infection on R. spp. It was considered applicable for early diagnosis of the “crooked-head disease” and breeding of healthy frogs for food and medicine.