Neutralization tests are of two types:
- Virus Neutralization test.
- Toxin Neutralization test.
Neutralization of a virus is defined as loss of infectivity through reaction of virus with specific antibody i.e. neutralization of viruses by their specific antibodies are called virus neutralization tests. Inoculation of viruses in cell cultures, eggs, and animals results in replication and growth of viruses. When virus-specific neutralizing antibodies are injected into these systems, replication and growth of viruses is inhibited. This forms the basis of virus neutralization test.
These are serological tests to detect presence and magnitude of functional systemic antibodies (Monoclonal and polyclonal) that prevent infectivity of a virus. E.g. viral hemagglutination inhibition test is virus neutralization test frequently used in diagnosis of viral infections such as influenza, mumps, and measles. If patient’s serum contains antibodies against certain viruses with the property of agglutinating red blood cells, these antibodies react with viruses and inhibits the agglutination of red blood cells.
Virus and serum are mixed under appropriate condition and then inoculated. The presence of unneutralized virus may be detected by reactions such as CPE, haemadsorption/hemagglutination, plaque formation, disease in animals. The loss of infectivity is bought about by interference of bound antibody with any one of the steps leading to release of viral genome into host cells.
It is important as it can be used to help determine prevalence of a disease, geographic dissemination of its virus, titer of antibody in an antiserum or principles underlying the immunologic factors of an infection. A known virus demonstrate its specific antibody, and known antibody reveals the identity of a virus.
Virus Neutralization Tests can be reversible neutralization or stable neutralization.
Neutralization process can be reversed by diluting Antibody-Antigen mixture within a short time of formation of Ag-Ab complexes (30 mins). It is thought that phenomenon is due to the interference with attachment of virions to cellular receptors e.g. attachment of hemagglutinin protein of influenza viruses to sialic acid which requires saturation of viral surface with antibodies.
Antigen-Antibody complexes usually gets stabilized with time (several hours) and the process cannot be reversed by dilution. Neither virions nor antibodies are permanently changed in stable neutralization, for the unchanged components can be recovered. The neutralized virus can be reactivated by proteolytic cleavage. It had been observed that neutralized virus can attach and that already attached virions can be neutralized.
Number of antibody molecules required for stable neutralization is considerably smaller than reversible neutralization (even a single antibody molecule can neutralize a virion). Such neutralization is generally produced by antibody molecules that establish contact with two antigenic sites on different monomers of a virion, greatly increasing the stability of the complexes. An example of stable neutralization is the neutralization of polioviruses, whereby, attachment of antibody to viral capsid stabilizes the capsid which inhibits uncoating and release of viral nucleic acid.
This virus-antibody complex can prevent viral infections in many ways. It may interfere with virion binding to receptors, block uptake into cells, prevent uncoating of genomes in endosomes, or cause aggregation of virus particles. Many enveloped viruses are lysed when antiviral antibodies and serum complement disrupt membranes. Antibodies can also neutralize viral infectivity by binding to cell surface receptors.