Production of Polyclonal Antibody against Mycoplasma ovipneumoniae in Rabbits Saroj Poudel Advisor: Dr. Jeffery J. Adamovicz Contributors: Dr. Gerard P.

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Presentation transcript:

Production of Polyclonal Antibody against Mycoplasma ovipneumoniae in Rabbits Saroj Poudel Advisor: Dr. Jeffery J. Adamovicz Contributors: Dr. Gerard P. Andrews and his lab Department of Molecular Biology, Department of Veterinary Science, Honors Senior Project, UW.

Introduction O Big Horn Sheep O Polyclonal antibodies in rabbits O Diagnostic assay

Overview Protein extraction  Swabs or pure culture Immunization ELISA Affinity chromatography

Attachment to epithelial cells Close association of attachment structure

Protein Extraction Specific protein from the pathogen or Membrane protein fractionation SDS Page BCA Lowry absorbance test

15 kd 25 kd 35 kd 50 kd 75 kd 100 kd 150 kd 225 kd Membrane Protein Prep

Immunization Adaptation Pre-bleed & Immunize Booster

Predicted chart for bleeding and immune response

ELISA Indirect Elisa  Primary antibody – rabbit antibody  Secondary antibody – Goat anti-rabbit antibody  TMB substrate

Affinity Chromatography Protein A chromatography Sepharose 4B chromatography

Conclusion Animal protocol Lab protocol Protein Membrane fractionation Bled Rabbits Immunized

Acknowledgements Dr. Jeffery J. Adamovicz Dr. Gerard P. Andrews and his lab members. Jake Berg

Supplements M. Pneumonia on Blood Agar Plate