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EQUINE VETERINARY EDUCATION / AE / july 2022


Scenario Red blood cells and virus a) Sample is seronegative b)


Red blood cells, virus and antibodies


Haemagglutination inhibition


Interaction


Haemagglutination Result


Sample is seropositive AA B B B


AA A


B


B


Fig 3: Haemagglutination inhibition assay; a) the covered red wells indicate haemagglutination (lattice of agglutinated cells), so there are no antibodies to the virus at those sample serum dilutions to neutralise the virus. b) red dot wells indicate no haemagglutination, so the test is positive at the specified dilution (the highest last dilution at which haemagglutination is prevented).


Disadvantages of CFT


• Anti-complimentary activity can interfere with the test, resulting in false-positive results


• Nonspecific due to cross-reactivity between EHV-1 and EHV-4


Indirect fluorescence antibody test


Fig 4: Serum radial haemolysis; top row, first two wells on left = negative samples, next four wells = positive samples. Middle row = empty wells. Bottom row, first three wells on left = mild positives, fourth well = empty, last two wells = positive controls.


whether the serum itself still fixes complement and prevents red cell lysis, mimicking what would happen if there were viral-specific antibodies present in the sera. If the serum demonstrates anti-complimentary activity, this must be considered when interpreting test results as they may be falsely elevated. CFT is used to test sample for the presence of equine herpes virus-1/-4 (EHV-1/-4) antibodies and this phenomenon of anti-complementary activity can be found in donkeys and occasionally in very sick horses (T.A. Hammond, personal communication).


Advantages of CFT


• A useful test for EHV-1/-4 as CF antibodies are not long lasting, even after vaccination


© 2021 EVJ Ltd


Indirect fluorescence antibody test (IFAT) involves sample serum being added to virus infected cells. If antibody is present in the sample, it binds with a specific antigen on the tissue. Fluorochrome-labelled secondary antibody conjugate is added, which binds to the already present antigen– antibody complex and releases fluorescence, seen on microscopy (Fig. 6). If there are no antibodies in the sample serum, there will be no fluorescence.


Advantages of IFAT


• Commonly used as a confirmatory test, for example samples being tested for Dourine that are CF positive will be confirmed by IFAT


Disadvantages of IFAT


• Test requires a longer time compared to other tests, for example CF and ELISA


• More expensive Virus neutralisation test


When virus is mixed with susceptible cells, it will infect them and when visualised under a microscope, a cytopathic


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