- Electro Chemiluminescence immunoassay
It is based on the technique of using streptavidin coated solid phase along with
ruthenium complex labeled antibodies for detection of analytes. The sample, the
biotinylated antibodies, antibodies labeled ruthenium complex, and streptavidin
coated micro particles are mixed into a reaction cup and incubated. The reaction
mixture is aspirated into an electrochemical measuring cell, and unbound conjugate
are washed away by TPA and a magnet. Electrical current is then used to excite the
ruthenium complex and initiate signal generation to detect the antigen-antibody
complexes in the sample.
- Enzyme immunoassay
This type of immunoassay employs the use of enzyme labeled antibodies for the detection
of antigen/antibodies in the sample. The enzyme catalyzes a color reaction when
exposed to the substrate.
- Immunoturbidometry
This measures the reduction in light transmission caused by particle formation.
- Electrophoresis and Immunofixation Electrophoresis
This method involves the separation of charged compounds based on their electrical
change. When a voltage is applied to a salt solution, an electric current is produced
by the flow of ions.
- Chromatography
It is a separation method based on different interaction of the specimen compounds
with the mobile phase and stationary phase as the compounds travel through a support
medium. Compounds interacting more strongly with stationary phase are retained longer
in the medium than those that from the mobile phase.
- Column chromatography
It is a type of chromatography in which the stationary phase, is placed in a vertical
glass (usually) column and the mobile phase, a liquid, is added to the top. This
liquid then flows down through the column (by either gravity or external pressure).
An equilibrium is established between the solute adsorbed on the adsorbent and the
eluting solvent flowing down through the column. Because the different components
in the mixture have different interactions with the stationary and mobile phases,
they will be carried along with the mobile phase to varying degrees and a separation
will be achieved. The individual component, or elutant, are collected as the solvent
drips from the bottom of the column. Column chromatography is separated into two
categories depending on how the solvent flows down the column. If the solvent is
allowed to flow down the column by gravity, or percolation, it is called gravity
column chromatography. If the solvent is forced down the column by positive air
pressure, it is called flash chromatography. Column chromatography is generally
used as a purification technique: it isolates desired compounds from a mixture.
- Nephlometry
It refers to the measurement of light scattered by a particulate solution. It measures
the concentration of a solution that contains particles too large for absorption
spectroscopy. It is important in measuring antigen-antibody reactions.
- Haemagglutination
The test uses the property of certain viruses to agglutinate red blood cells. This
occurs due to the binding of haemagglutinin part of the viral protein to receptors
on the membrane of red blood cells. This linking results in visible macroscopic
clumping, known as haemagglutination.
- Latex agglutination
It is a type of passive agglutination reaction in which polystyrene latex particles
are attached to the surface of a soluble antigen. This step converts a precipitation
reaction into a agglutination reaction. The latter being more convenient and sensitive
for the detection of antibodies.
- Radial Immunodiffusion
It is a type of precipitation reaction in which the antibody/antiserum is incorporated
in agar gel poured on a flat surface (slide or Petri dish). The antigen is added
to the well cut on the surface of the gel. The antigen diffuses radially from the
well and forms ring shaped bands of precipitation concentrically around the well.
The diameter of the halo gives an estimate of the concentration of the antigen.
- Indirect Immunoflorescence
Immunoflorescence assays use fluorescent conjugated antibodies to locate antigens
in samples and tissues. In indirect Immunoflorescence technique, the antigen is
first mixed with the antibody and then treated with fluorescent conjugated antiglobulin
serum. On examination under UV illumination, antigen-antibody complexes are seen
as fluorescing objects against a dark background.
- Fluorescence Polarization Immunoassay
In this assay, fluorescein-labelled drug competes with unlabelled drug for antibody.
On excitation of the sample with plane polarized light (490 nm). Fluorescein emits
plane polarized light (520 nm). Small, free drug-fluorescein, rotates faster leading
to less emission while larger, antibody- drug- fluorescein, rotates slower and produces
more emission. Thus, more drug in the sample: less fluorescein labeled drug bound
to antibody lower emission of plane polarized light.
- Flourescent Microscopy
The excitatory light is passed from above (or, for inverted microscopes, from below),
through the objective lens and then onto the specimen instead of passing it first
through the specimen. The fluorescence in the specimen gives rise to emitted light
which is focused to the detector by the same objective that is used for the excitation.
Since most of the excitatory light is transmitted through the specimen, only reflected
excitatory light reaches the objective together with the emitted light and this
method therefore gives an improved signal to noise ratio. An additional filter between
the objective and the detector can filter out the remaining excitation light from
fluorescent light.