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Measuring Antibody/Antigen Reactions

Measuring Antibody/Antigen Reactions Binding of antibody to antigen by itself is invisible: how can it be detected & quantitated ?.

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Measuring Antibody/Antigen Reactions

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  1. Measuring Antibody/Antigen Reactions Binding of antibody to antigen by itself is invisible: how can it be detected & quantitated? One visible consequence of Ab/Ag binding may be precipitation, and one technique for detecting Ag/Ab reactions by precipitation in a semi-solid gel is the Ouchterlony assay.

  2. Ouchterlony Analysis: 2-Dimensional Precipitation in Agarose Several distinct patterns of reactivity are shown above – the following animations illustrate how some of these patterns may be formed.

  3. Ouchterlony: Double Diffusion in Agarose Simple Precipitin Line Antigen (HSA) and antibody (Ab) each begin to diffuse radially away from their wells. Where they meet, a line of precipitation begins to develop. Remember that Ab/Ag binding is reversible; this line could move by dissolving along one edge and re-precipitating along the other, depending on the relative concentrations of Ab and Ag… Ab = rabbit anti-HSA

  4. Ouchterlony: Double Diffusion in Agarose Reaction of Partial Identity X,Y X aY aX,aY The “αX+X” line (formed between “Ab” and “BSA”) cannot extend any further upward because the “αX” antibody cannot cross the other precipitin line (since that line also contains epitope “X”) Ab = rabbit anti-HSA

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