A) Nucleic acid amplification

Most commonly, PCR (polymerase chain reaction) is used for both DNA and RNA viruses. For RNA viruses, an initial reverse transcription step which converts RNA to DNA is used. PCR is highly sensitive and relatively rapid.
Other variations of nucleic acid amplification are used such as LCR (ligase chain reaction) for Chlamydia trachomatis, NABSA (nucleic acid sequence based amplification) HIV viral load etc. HIV viral load allows for monitoring disease progression and response to treatment.


B) Serology.

Two main methods available for antibody detection in blood

1. ELISA (enzyme-linked immunosorbent assay) :
ELISA detects antibodies (IgM, IgG and IgA). IgM is detectable immediately after the onset of illness. IgG antibodies, which appear later, indicate past infection and immunity in most instances. ELISA can be adapted to detect antigens (e.g. HBsAg).

2. CFT (complement fixation test) :
Antigen is added to patient’s serum; if specific antibody is present, then a complex forms which binds complement, so complement is no longer free to lyse indicator red blood cells, added to well. The result is given as a titre which indicates the amount of antibody present. A rise in titre between an acute and convalescent serum (10-14 days after onset of illness) is required to make a diagnosis.


C) Direct antigen detection by immunofluorescence

Certain specimens with lots of cells such as nasopharyngeal aspirates (NPA) and bronchoalveolar lavages (BAL) can be stained directly with monoclonal antibodies prepared against specific pathogens such as influenza virus, respiratory syncytial virus (RSV), adenovirus and Pneumocystis carinii. A good quality specimen is needed!


D) Electron microscopy (EM)

This requires a specimen where high concentration of virus in samples can be expected, such as diarrhoeaic stool and vesicle fluids. EM has become a reference technique.


E) Virus isolation

Samples are inoculated onto several different types of cells (e.g. derived from kidney or lung) selected for the types of viruses expected in the clinical sample. The cultures are inspected for evidence of cytopathic effect (effect of the virus on the cell). This technique depends on the presence of live virus. So the use of correct viral transport medium, the time the sample takes to reach the lab, and correct storage of samples prior to transfer are all important factors. Not all viruses can be grown in cell cultures.


Monolayer of normal uninfected cells

Monolayer of cells showing viral cytopathic effect

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