METHODOLOGIC PROBLEMS IN ESTABLISHING NORMAL VALUES FOR IGG SUBCLASS CONCENTRATIONS IN A PEDIATRIC POPULATION - COMPARISON OF RADIAL IMMUNODIFFUSION AND ELISA METHODS

The aim of this study was to establish an enzyme-linked immunosorbent assay (ELISA) to measure IgG subclasses by means of monoclonal antibod ies. The distribution of IgG subclass protein concentrations in sera f rom 227 healthy Danish children and 90 adults was measured. Furthermor e, this newly established ELISA was compared with different assay syst ems for determination of IgG subclasses: two radial immunodiffusion me thods (RID), one using polyclonal and one using monoclonal antibodies, as well as a commercially available ELISA kit. There was good agreeme nt of results obtained by the different methods of measuring IgG3 and IgG4 concentrations. There was good correlation between results obtain ed by both RID methods. Despite good correlation between the assays, t he ELISA kit showed higher levels of IgG1 in all investigated sera, an d the ELISA kit showed no correlation with the other methods, when IgG 2 was measured. Analysis of the normal ranges measured by ELISA develo ped in our laboratory and by RID with polyclonal antibodies showed tha t the levels obtained by RID were higher than those obtained by our EL ISA in sera with low levels of both IgG1 and IgG2, and lower in sera w ith high concentrations of these two immunoglobulins. Our results emph asize the importance of establishing age-related normal limits for any novel assay measuring IgG subclass concentrations.