This technique is used to indirectly detect the presence of complements
bound to the antigen-antibody complexes using no lysis of sensitized red
blood cells as an indicator.
High group specificity
Antibodies disappear relatively quickly.
Screening test for infection
Hemagglutination inhibition (HI)
For viruses with hemagglutination activity, hemagglutination-inhibiting
antibodies can be determined.
High specificity for type
The antibody concentration increases from the early phase
of infection and maintained thereafter.
Fluorescent antibody (FA)
Reaction between a specific antigen in infected cells and the corresponding
antibody is demonstrated using fluorescence-labeled antibodies.
Antibody fractionation is available.
Neutralization test (NT)
Active viruses are neutralized by antibodies to demonstrate protective antibody responses.
High specificity for type
Enzyme immunoassay (EIA)
Solid-phase antigens are reacted with enzyme-labeled antibodies to
demonstrate the presence of the antigen of interest.
Antibody fractionation is available.
Quantitative data
High sensitivity compared with other techniques
Passive (particle)
agglutination (PA)
Using gelatin particles coated with viral antigens, antigen-antibody
reaction is carried out. The existence of the antigen is determined based
on appearance of clots.
High sensitivity
Western blotting (WB)
This technique is used to detect specific antibodies that specifically react
with antigen protein bands that are separated and transferred to membrane.
High specificity
Identification test
Properties of detected antibodies
Properties
Kinetics
Antiviral antibody activity
Complement binding capacity
Placental transfer
Antibody
IgM
IgM antibodies are produced at an early stage after onset of
infection and disappear in a short period.
+
+
-
IgG
IgG antibodies are produced after IgM antibodies and
persist longer than IgM antibodies with a gradual decrease.
+
+
+
IgA
IgA antibodies are produced a little later than
IgM antibodies and persist longer than IgM antibodies.
+
-
-
Interpretation of antibody titer and the significance of a paired serum test
The notion of the normal value does not exist in serum antibody titer against the infecting virus.
Detecting an antibody that is produced after virus infection only retrospectively indicates
the past infection of the virus and does not necessarily reflect the current condition.
Antibody titer against a virus shows a pattern that is high shortly after the infection then declines,
but it is often impossible to determine whether there was infection in the immediate past only with
the level of single serum antibody titer.
It is necessary to select a test method according to the purpose by understanding patterns
of antibody response after virus infection, features of each test method,
and the significance of the test.
A 4-fold or larger rise in the antibody titer of the paired serum, one from the acute phase
(early post-onset phase) and the other from the convalescent phase (14–21 days after onset),
is judged as significant and infection of the virus is presumed. However,
a rise in antibody titer in the case where gamma globulin was administered for
treatment is not necessarily considered significant.
Guide for selecting a test method according to the purpose
It is necessary to understand the features of each test method and select a test according to the purpose.
In the case of natural infection, it is helpful to detect IgM antibodies that respond in
the early phase of infection and observe a rise in antibody titer of the paired serum.
Also, a test for lgG antibodies by EIA is helpful in determining the past history and vaccine effect.
Natural infection
Past history
Vaccine effect
Measles
NT、EIA (IgM) (IgG)
NT、EIA (IgG)
NT、EIA (IgG)
Rubella
HI、EIA (IgM) (IgG)
HI、EIA (IgG)
HI、EIA (IgG)
Mumps
CF、HI、NT、EIA (IgM) (IgG)
EIA (IgG)
NT、EIA (IgG)
Varicella
CF、EIA (IgM) (IgG)
EIA (IgG)
EIA (IgG)
Japanese encephalitis
HI、CF
HI
Influenza
CF、HI
HI
How to order the test and interpret the test result
Because measured values vary depending on the disease condition, please make sure to order serums of the acute phase
(early post-onset phase) and the convalescent phase (2–3 weeks after onset) in pairs.
If measurements, taken simultaneously for the acute phase and the convalescent phase,
show a four-fold or larger rise in antibody titer, the result is considered serologically significant.
Characteristics of virus antigen test
Methodology
Principles
Characteristics
Shell vial method
Culture virus sensitive cells on a slide glass in a shell vial (drum-shaped container),
and inoculate specimen into the vial. After centrifugation,
incubate the mixture for 24 to 48 hours.
Then, with the fluorescent antibody (FA) technique using viral antigen-specific antibodies,
detect virus-specific antigens that have been cultured.
Infectious viruses can be identified in
relatively shorter time than virus isolation and culture.
Enzyme immunoassay (EIA)
After reaction between viral antigens and specific antibodies,
the antigens are detected using enzyme reaction.
In the direct method, specific antibodies are directly labeled with enzyme,
whereas in the indirect method, secondary antibodies are labeled with enzyme.
High sensitivity
Fluorescent antibody（FA)
method
After reaction between viral antigens and specific antibodies,
the antigens are detected using fluorochrome. In the direct method,
specific antibodies are directly labeled with a fluorochrome,
whereas in the indirect method, secondary antibodies are labeled with a fluorochrome.
High specificity
Polymerase chain reaction (PCR)
Heat denatured single-stranded DNA is bound to a target primer
(DNA fragments of 20 to 30 bases each complementary to a specific target DNA region to be amplified).
DNA polymerase activity catalyzes the DNA synthesis.
By repeating the cycles, the target DNA sequence is amplified in an exponential manner.
High sensitivity and specificity
Southern blot hybridization
After restriction enzyme digestion of DNA,
the DNA is isolated and denatured to a single-stranded form with agarose electrophoresis.
The single-stranded DNA is transferred to a membrane and
hybridized with a labeled probe to detect the target gene.
Analysis of abnormal changes in DNA in a quantitative and qualitative manner
