Characterization of immunochemical reaction for human growth hormone with its monoclonal antibody by perfusion protein G affinity chromatography and capillary zone electrophoresis.
An extremely rapid assay technique for antibody-antigen interaction using human growth hormone and its monoclonal antibody as an example has been developed by utilizing Protein G bound to perfusion chromatography matrices. The antibody and antigen were mixed and incubated at different molar ratios by keeping the concentration of antibody constant. The mixture of antibody and antigen solution was then injected onto the Protein G column. The complex of antibody and antigen formed in the sample solution was retained on the Protein G column as was the free antibody. The peak-area and height of the retained complex and antibody linearly increased with the molar concentration of the antigen when it was not in excess of the corresponding stoichiometric amount of antibody, however, those of the retained peak were constant when the concentration of antigen was in excess of that of the antibody. The validity of this method was confirmed by the results of capillary zone electrophoresis. The method developed cannot only be used to determine the biological activity between antibody and its antigen quickly, but also to determine the stoichiometry of immunological reactions between the antibody and antigen when a stable complex of them can be formed.
Zou
H
H
National Chromatographic R. and A. Center, Chinese Academy of Sciences, Dalian, China.
Zhang
Y
Y
Lu
P
P
Krull
I S
IS
D000911:Antibodies, Monoclonal / Q000737:chemistry*; D002846:Chromatography, Affinity; D019075:Electrophoresis, Capillary; D013006:Growth Hormone / Q000737:chemistry*; D006801:Humans; D007120:Immunochemistry; D009419:Nerve Tissue Proteins / Q000737:chemistry*; D013056:Spectrophotometry, Ultraviolet
D000911:Antibodies, Monoclonal; C029152:G-substrate; D009419:Nerve Tissue Proteins; D013006:Growth Hormone