Relative avidities using elution with a single dilution of the chaotrope were expressed in terms of the percent reduction in IgG concentration compared to the concentration in the absence of the chaotrope (2, 11)

Relative avidities using elution with a single dilution of the chaotrope were expressed in terms of the percent reduction in IgG concentration compared to the concentration in the absence of the chaotrope (2, 11). Data analysis. lower than the correlation between the two elution methods (= 0.84). We found that the serum dilution, the heterogeneity of the antibody population, and the concentration of the chaotrope were the primary variables affecting avidity determinations. In this study, we present multiple analysis methods depending on the methodology used. We also present the factors that affect the analysis of avidity determinations given the polyclonal nature of human sera. This experimental approach should benefit the evaluation of similar antibodies induced by other bacterial polysaccharide vaccines. In the United States there has been a dramatic decrease in the incidence of Sec-O-Glucosylhamaudol type b (Hib) disease since the introduction of highly efficacious vaccines in the 1980s (6-8). This success is attributed to the widespread vaccine coverage across the country and to the capacity of polyribosylribitol phosphate (PRP) protein conjugates to elicit long-term protection that can be recalled upon exposure or boosting with a subsequent dose (3, 5, 10, 16). This recall of the established memory has been observed even when a reduced dose of Sec-O-Glucosylhamaudol vaccine is used (9). Vaccination elicits memory B- and T-cell clones. With time, the surviving clones tend to be those producing antibodies of higher avidity. Recall of highly avid antibodies by the vaccine antigen would indicate that efficient memory was established. Goldblatt et al. have shown that this is the case with Hib and that antibody avidity can be a marker for the presence of immunological memory (11). However, the long-term benefit of immunological memory as measured by avidity estimates may require additional investigation, especially when only vaccination regimens with three doses are used for young children, as in the case of the meningococcal serogroup C conjugate vaccine (23). Antibody affinity can be defined as the strength of the binding of a single antibody type (a homogeneous antibody, such as a monoclonal antibody) and a single antigenic target (hapten). In this single-epitope interaction, the affinity constant is the amount of complexed antigen-antibody at equilibrium (13). In human serum the antibody population is heterogeneous (polyclonal in nature), and determination of antibody affinity is not possible. However, adaptations of this affinity concept have been devised, and determinations of the stability of the antigen-antibody interactions in a mixed population of antibodies have been termed antibody avidity determinations (13). ANGPT1 Chaotropic agents such as urea or thiocyanate have been preferred for the determination of the average estimate of antibody avidity. In the laboratory, antibody avidity can be estimated by using a variety of methods. Therefore, antibody avidity estimates will differ according Sec-O-Glucosylhamaudol to the methodology used. Each methodology has its own limitations, and it has been difficult to compare the results of one study with those of another given the diversity of approaches to experimentation and analysis. In this investigation, we compare three experimental approaches that represent some of the methodologies used for the determination of anti-PRP antibody avidity (11, 20) and the knowledge gained from other avidity methods used for (2, 24). We present the limitations of each approach and the preferred method of analysis for the data obtained. Similar evaluation of antibody avidities elicited by other bacterial polysaccharide vaccines should benefit from.