ELISA Tips on Incubation

1. Rotation of plates while incubating reagents.
2. In certain ELISA systems, the plates are rotated during incubation for better antigenantibody reaction.
3. The effect of rotating plates is to mix the reactants completely during the incubation step.
4. Since the solid-phase limits the surface area of the absorbed reactant, the mixing ensures that, potentially reactive molecules are continuously coming into contact with the solid-phase.
5. During stationary incubation, mixing only takes place because of diffusion of reagents.
6. Thus, to allow maximum reaction from reagents in stationary conditions, greater times of incubation may be required, than if they are rotated.
7. Rotation also allow ELISA to be performed independent of temperature conditions.
8. The interaction of antigen & antibodies relies on their closeness, and the kinetic energy provided to the system, which is encouraged with the mixing during rotation.
9. Stationary incubation relies on the diffusion of molecules & thus is dependent on temperature.