Serum heated to 56 for 30 minutes to destroy non-specific inhibition of viral infectivity. A dilution of serum is mixed with a constant amount of virus, mixture allowed to incubate at room temperature for 60 minutes, and decrease in plaques measured . End point taken is highest dilution to give inhibition. Note higher the number the greater the no. of antibodies. Neutralizing antibodies show very little cross reactivity, even within the same species.
Lets look at neutralization in a little more detail.
If influenza virus is mixed with antibody at 0 C and a sample of the mixture is added to an assay system half an hour later without dilution a decrease in the viral titer in comparison with untreated virus may be seen, but if the neutralization mixture is diluted by a large factor before the assay, the original titer is restored. There is a freely reversible equilibrium between dissociation and reformation of the Ab-virion complex.decreasing the concentration of the reactants diminishes the rate of complex formation but not the rate of dissociation. With time the antibody virus complexes become more stable. If the assay is made several hours after the virus has been mixed with the antibody, neutralization persists even after dilution. Whereas reversible complexes form readily at 0 c,with little temperature dependence,the formation of irreversible complexes occur at 37, suggesting the need for configurational changes.