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Automated nephelometric immunoassay (ANIA) I. Importance of antibody affinity

Authors
Journal
Journal of Immunological Methods
0022-1759
Publisher
Elsevier
Publication Date
Volume
5
Issue
2
Identifiers
DOI: 10.1016/0022-1759(74)90006-4

Abstract

Abstract The reason why certain antisera displaying good precipitating properties in gels may be quite inefficient in automated nephelometric immunoassays has been sought. Antibodies against dinitrophenyl groups, gamma chains, or haptoglobin were eluted by increasing concentrations of ammonium thiocyanate from their respective antigens coupled to Sepharose. It was found that those antibodies that required the highest concentrations of thiocyanate to dissociate them from their antigen, were also those that formed aggregates with the most elevated nephelometric effect. Antibodies against dinitrophenyl groups were also eluted increasingconcentrations of the specific hapten. After dissociation from the latter, these antibodies displayed a nephelometric effect clearly dependent on their affinity. By increasing the concentration of antibodies (isolated on immunoadsorbents), and lowering the reaction temperature, it was possible to compensate for the lack of affinity of certain antisera and to make them suitable for automated nephelometric immunoassay.

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