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The use of antibodies is widespread in many areas including in-vivo and in-vitro diagnostics, quantitative analysis in research laboratories and as therapeutic substances. Since the methods for generation of antibodies has improved and regularly results in high-affinity interactions, the standard assays used for quantification of the interaction properties should be revisited because they do not necessarily produce accurate results. Here we show that in several cases, the affinity determination of strongly binding antibodies will be inherently difficult when using standard procedures, due to impractically long incubation times. Real-time kinetic analysis is often the only realistic alternative for affinity determination.

Antibody-antigen interactions: What is the required time to equilibrium?