Methods of Laboratory Diagnosis
Laboratory diagnosis of viral infections can be carried out by many methods. These methods include
(a) demonstration of virus-induced cytopathic effects (CPEs) in the cells,
(b) direct detection of viruses,
(c) virus isolation and viral assays,
(d) detection of viral proteins and other enzymes,
(e) detection of viral genome, and
(f) viral serology (Fig. 53-1).
Viral serology is based on detection of specific viral antibod-ies in serum of the infected human host. A wide number of serological tests are used for demonstration of specific viral antibodies in patient’s sera. These include hemagglutination inhibition (HI) test, neutralization test (NT), indirect fluores-cent antibody (IFA) test, ELISA, RIA, latex agglutination test (LAT), and Western blot.
The detection of virus-specific immunoglobulin (IgM) antibody in serum indicates a recent primary infection. This is because IgM antibodies appear in the serum during first 2 or 3 weeks of primary infection. A fourfold increase in the antibody titer between the serum collected during the acute phase of the disease and during the convalescent phase, 2–3 weeks after acute phase, is suggestive of seroconversion. An anamnestic or secondary antibody response occurs during reinfection or recurrence of viral infection later in life. The serum antibody titer usually remains high in individuals who suffer frequent recurrence of the disease, such as herpes virus.
· HI test is used for detection of viruses that agglutinate red blood cells of chickens, guinea pigs, human, or other mammals. Antibodies present in serum prevent viruses to bind and to agglutinate the new blood cells. The HI test is used for diagnosis of infections caused by orthomyxoviruses.
· NT is based on inhibition of infection by the antibody and that of CPEs of the viruses in tissue culture cells. The neu-tralization antibody response is usually virus and strain spe-cific. The highest dilution of serum preventing infectivity is 50% of virus. Serum mixture tested is considered as the titer of the test.
· ELISA is most commonly used for screening of blood samples for hepatitis B and C viruses and HIV.
· IFA, RIA, and LAT are also used for diagnosis of many common viral infections.
· Western blot is most commonly used to confirm infection caused by HIV, initially diagnosed by ELISA.
The limitations of serological tests in viral diseases are the following:
· The presence of antiviral antibody in serum only indicates infection but cannot determine whether it is recent or old. Demonstration of IgM antibodies or demonstration of a fourfold increase in the antibody titer between acute and convalescent sera indicates only recent infection.
· The serological tests may be associated with false-positive or false-negative reactions. The serological cross-reaction may occur between different viruses, giving rise to false-positive reactions. Formation of immune complexes in serum may give rise to false-negative reaction as observed in viral infection caused by hepatitis B virus.