(A) Monoclonal-antibody production consists of sequential procedures: immunogen production, immunization and boosting, cell fusion, clonal selection, ascites production and antibody purification; overall, monoclonal antibody production takes approximately 7 months. (B) Antibody titer in the blood and the level of immunization was regularly assessed by direct ELISA of crude serum on immunogen coated plates. After the second and third immunization, 10 ul of blood of immunized mice was tested. (C) Supernatant of each clone was used for the western blot analysis to detect SAA protein, used in the immunization. Clones were selected by the results of western blot analysis. (D) Pure antibodies were purified from ascites by passage through IgG immunoaffinity columns. The CBB staining results show that highly purified IgG against hu-SAA. (E) Purified anti-SAA antibody sensitivity and specificity was tested by western blot analysis on the synthetic SAA.
