Abstract
Antigen Used for Immunization
Monoclonal antibody against human serum albumin was produced by hybridoma technology with high purity.
Method of Immunization
Human serum albumin was used as the immunogen. The BALB/c mice were immunized by subcutaneous injection of 0.1 mL (containing 50 μg human serum albumin) immunogen. Thirty μg of antigen prepared in FIA was used for booster immunization every 5 days for 5 months.(1,2)
Parental Cell Line Used for Fusion
SP2/0 mouse myeloma cells.
Selection and Cloning Procedure
Hybridoma-producing monoclonal antibodies to HSA were grown in a 20% FCS containing medium supplemented with 1% hypoxanthine-aminopterin-thymidine (HAT) for 3 weeks. The supernatants were tested for reactivity to HSA by ELISA. As a specific and non-specific capture antigen, HSA and hemoglobin, respectively. were used. Positive hybridomas were subsequently cloned two times by limiting dilution method.(2,3)
Heavy and Light Chains of Immunoglobulin
The isotype was determined using the Sigma ImmunoType Kit (St. Louis, MO). This clone was found to be of IgG class with kappa light chain.
Specificity
Monoclonal anti-human serum albumin reacts specifically against human albumin using competitive ELISA assay based on inhibition of antibody binding to immobilized antigen. The antibody was specific for human serum albumin, and low cross-reaction was observed with bovine serum albumin (BSA), hemoglobulin, and human immunoglobulin.(2,3)
Specific Antigen Identified
N/A