Measurement of Specific IgG Antibody with Protein-A Sepharose CL-4B

Abstract

Evaluation of the method for measurement of specific IgG by using protein-A Sepharose CL-4B was carried out(for the puropose of clinical application). The results obtained are as follows. 1) conjugation of IgG to protein-A Sepharose CL-4B is specific. This is confirmed by both affinity chromatography and immunoelectrophoresis method. 2) The maximum value of the specific IgG was observed when 50μl of serum was added to one ml of the protein-A Sepharose CL-4B comples. 3) Two inhibition tests were performed in order to determine whether the specific antibodies are truly measured or not. One was incubation with unlabeled insulin antigen and the other was incubation with unlabeled mite antigen. The results were compared respectively with those obtained with those with by the atandard methods. The results obtained with the inhibition tests were almost identical with those calculate theoretically which indicate the effect of inhibition. 4) Washing after incubation with <125>^I labeled antigen is necessary for 6 times. 5) a) The specific IgG antibodies to insulin are measured twice on twenty six sera. The correlation coefficient between the results of two measurements is 0.9885 is highly significant. (p<0.0001) b) Evaluation of technical error in measurement was carried out by doing duplicate tests on 24 samples of IgG to mite. Difference was within 10% and 5.9% in average. 6) Significant correlation coefficient was observed specific IgG and IgE antibody to egg white albumin (r=0.35, t=2.56, p<0.02). But no significant correlation was found in case of β-lactoglobulin.