Figure 24:Techniques that require purified antibodies. Technique Antibodies use Antibody type Best source Comments Cell Strain Direct Local Anti-antigen Polyclonal-Monoclonal Prepare yourself Indirect local Anti-antibody Polyclonal Commercial Immunoassay Direct detection Anti-antigen Monoclonal-Polyclonal Prepare yourself Indirect detection Anti-antibody Polyclonal Commercial Immunoblot Direct detection Anti-antigen Polyclonal-Monoclonal Prepare yourself Indirect detection Anti-antibody Polyclonal-Monoclonal Commercial Affinity Purification Anti-antigen Monoclonal Prepare yourself Figure 25:Sources for purifying antibodies. Cor Source Ab Conc.Total Type Ab eifie Posible Contaminating Purity of specif (mg/ml) (mg/ml) Serum Polyclonal 10 1%to 10%Other Serum Ab 10% 0.05%Calf Ab 95% Tissue Culture Monoclonal 0.05 0.05%None 95% Ascities Monoclonal 1-10 90% Mouse Ab 90% Quantity and quality of purified antibodies During the purification of antibodies,several variables need to be monitored.These include the purity,the amount of protein,the antigen binding activity of the antibody,and specific activity (Figure 26). Purity At any stage,the simplest method to determine the Figure 24: Techniques that require purified antibodies. Technique Antibodies use Antibody type Best source Comments Cell Strain Direct Local Anti-antigen Polyclonal-Monoclonal Prepare yourself Indirect local Anti-antibody Polyclonal Commercial Immunoassay Direct detection Anti-antigen Monoclonal-Polyclonal Prepare yourself Indirect detection Anti-antibody Polyclonal Commercial Immunoblot Direct detection Anti-antigen Polyclonal-Monoclonal Prepare yourself Indirect detection Anti-antibody Polyclonal-Monoclonal Commercial Affinity Purification Anti-antigen Monoclonal Prepare yourself Figure 25: Sources for purifying antibodies. Source Conc. Ab Type Conc.Total Ab Specific Ab Possible Contaminating Ab Purity of Specific Ab (mg/ml) (mg/ml) Serum Polyclonal 10 1% to 10% Other Serum Ab 10% Tissue Culture PBS 10% Monoclonal 1 0.05% Calf Ab 95% Tissue Culture Monoclonal 0.05 0.05% None 95% Ascities Monoclonal 1-10 90% Mouse Ab 90% Quantity and quality of purified antibodies During the purification of antibodies, several variables need to be monitored. These include the purity, the amount of protein, the antigen binding activity of the antibody, and specific activity (Figure 26). Purity At any stage, the simplest method to determine the