1. Antibody Detection

2. Part II

3. Detection of Antigens Western Blotting “Dot-Blot Assay” Positive Control Primary antibody (IgY) Lysed E. Coli Cells Lysed Salmonella Cells Nitrocellulose Membrane Has a high affinity to all proteins 1 Binding Ag (antigen) onto membrane. (E. Coli & Salmonella) Pre-lab & Lab Protocol #1

5. 2. Blocking with 3% BSA. In 1% PBS (Phosphate buffer solution) Lab Protocol #2 - 3 Mrs. Bielic say’s: BSA is a Large inert’t’t’ Protein molecule

7. STOPSWITCH TO PART I

8. 3. Application of Primary Antibody (IgY) Your Antibodies you isolated form chicken egg Lab Protocol: #4

9. 4. Rinse non-specifically bound Primary Antibodies with PSB (Phosphate buffer solution) Lab Protocol # 5 - 7

10. 5. Application of Secondary antibody (HRP-Horseradish Peroxidase Conjugated, goat anti – IgY) Lab Protocol # 8 - 9

11. 6. Rinse non-specifically bound Secondary Antibodies with PSB (Phosphate buffer solution) Lab Protocol # 10 -12

12. 7. Develop with TMB (HRP Substrate) Lab Protocol # 13 -16

13. BSA Salmonella E. coli Primary Ab Secondary Ab TMB BSA