Test methods

The following test methods are carried out in our laboratory:

Test methods

Fluorescent Antibody Virus Neutralization (FAVN) Assay
FAVN assay is a serum neutralization assay and serves to the determination of antibodies against rabies virus. Test sera are diluted and incubated with a constant amount of test virus, finally cells susceptible for the virus are added. NEutralizing antibodies in the test sera prevent a cytopathic effect in the used indicator cells. Virus reproduction is detected by direct immunofluorescence staining as rabies virus is a non cytopathogenic virus. Neutralization titer is calculated using the Spearman–Kärber method and indicated as international units (IU) compared to a determinated standard. This assay is used for vaccine efficiency control or as a prescribed test in accordance to travel purposes with dogs, cats and ferrets. More information regarding the latter you find here.
Serum neutralization assay
Virus neutralization can be used to detect and quantify neutralizing antibodies in serum samples. Therefore cytopathogenic virus strains are used that lead to visible cytopathic effect. If non cytopathogenic viruses are used (e.g. rabies virus) the cpe has to be made visible by immunofluorescence staining.
For this assay sera are diluted and incubated with a defined amount of serum for a certain time period. After addition of an indicator system (culture cells) the test is incubated for some days (usually 3-5 days, for some viruses 8 days can be necessary).
Using statistics the neutralization titer of the respective serum is calculated. A statement of the course of infection can only be made if a paired serum sample is tested. This assay can't differentiate between antibodies due to a vaccination and such due to a natural infection (even if a marker vaccine is used). In most cases a titer can't be related to a certain protectivity against a virus.
Advantages: high sensitivity and specificity.
Diasadvantages: time, material and staff consuming method; assay difficult to be standardized between laboratories due to different virus strains and cell culture systems used.
Indirect immunofluorescence assay
Indirect immunofluorescence can be used to detect antibodies against virus antigens from patient sera. Test cells infected with a known virus isolate are incubated with the serum in question and the binding of antibodies from the serum is detected using a fluorochrome-labeled secondary antibody. Indirect immunofluorescence is used e.g. to detect antibodies against the feline coronavirus (FCoV, including pathogen of feline infectious peritonitis (FIP)), against the Borna disease virus (BoDV) of mammals and the avian Bornavirus (mainly PaBV).
Advantages: fast process with high sensitivity.
Disadvantages: high equipment expenditure, unspecific reactions possible (depending on the test material and the quality of the antibodies).

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