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Immunofixation permits the detection and typing of
Immunofixation permits the detection and typing of monoclonal antibodies
A monoclonal antibody (mAb, more rarely called moAb) is an antibody produced from a Lineage (evolution), cell lineage made by cloning a unique white blood cell. All subsequent antibodies derived this way trace back to a unique parent cell.
Mon ...
or immunoglobulins
An antibody (Ab) or immunoglobulin (Ig) is a large, Y-shaped protein belonging to the immunoglobulin superfamily which is used by the immune system to identify and neutralize antigens such as bacteria and viruses, including those that cause di ...
in serum
Serum may refer to:
Biology and pharmacology
*Serum (blood), plasma from which the clotting proteins have been removed
**Antiserum, blood serum with specific antibodies for passive immunity
*Serous fluid, any clear bodily fluid
Places
*Serum, Ind ...
or urine
Urine is a liquid by-product of metabolism in humans and many other animals. In placental mammals, urine flows from the Kidney (vertebrates), kidneys through the ureters to the urinary bladder and exits the urethra through the penile meatus (mal ...
. It is of great importance for the diagnosis and monitoring of certain blood related diseases such as myeloma
Multiple myeloma (MM), also known as plasma cell myeloma and simply myeloma, is a cancer of plasma cells, a type of white blood cell that normally produces antibodies. Often, no symptoms are noticed initially. As it progresses, bone pain, a ...
.
Principle
The method detects byprecipitation
In meteorology, precipitation is any product of the condensation of atmospheric water vapor that falls from clouds due to gravitational pull. The main forms of precipitation include drizzle, rain, rain and snow mixed ("sleet" in Commonwe ...
: when a soluble antigen
In immunology, an antigen (Ag) is a molecule, moiety, foreign particulate matter, or an allergen, such as pollen, that can bind to a specific antibody or T-cell receptor. The presence of antigens in the body may trigger an immune response.
...
(Ag) is brought in contact with the corresponding antibody
An antibody (Ab) or immunoglobulin (Ig) is a large, Y-shaped protein belonging to the immunoglobulin superfamily which is used by the immune system to identify and neutralize antigens such as pathogenic bacteria, bacteria and viruses, includin ...
, precipitation occurs, which may be visible with the naked eye or microscope.
Immunofixation first separates antibodies
An antibody (Ab) or immunoglobulin (Ig) is a large, Y-shaped protein belonging to the immunoglobulin superfamily which is used by the immune system to identify and neutralize antigens such as bacteria and viruses, including those that caus ...
in a mixture as a function of their specific electrophoretic
Electrophoresis is the motion of charged dispersed particles or dissolved charged molecules relative to a fluid under the influence of a spatially uniform electric field. As a rule, these are zwitterions with a positive or negative net ch ...
mobility. For the purpose of identification, antisera are used that are specific for the targeted antibodies.
Specifically, immunofixation allows the detection of monoclonal antibodies representative of diseases such as myeloma or Waldenström macroglobulinemia
Waldenström macroglobulinemia ( , ) is a type of cancer affecting two types of white blood cells: lymphoplasmacytoid cells and plasma cells (both B cells). It is characterized by having high levels of a circulating antibody, immunoglobulin M (I ...
.
Technique
The technique consists of depositing a serum (or urine which has been previously concentrated) sample on a gel. After application of an electric current that allows the separation ofproteins
Proteins are large biomolecules and macromolecules that comprise one or more long chains of amino acid residues. Proteins perform a vast array of functions within organisms, including catalysing metabolic reactions, DNA replication, re ...
according to their size, antibodies specific for each type of immunoglobulin
An antibody (Ab) or immunoglobulin (Ig) is a large, Y-shaped protein belonging to the immunoglobulin superfamily which is used by the immune system to identify and neutralize antigens such as pathogenic bacteria, bacteria and viruses, includin ...
are laid upon the gel. It thus appears to be more or less narrow bands on the gel, which are at different immunoglobulins.
Immunofixation as immunoelectrophoresis, takes place in two steps:
* The first step is identical for both techniques. It consists in depositing the immunoglobulins contained in the serum or urine on a gel and then separating the immunoglobulins according to their electrophoretic mobility by making them migrate under the effect of an electric field. This migration depends on the mass and charge of the antigen. Once the immunoglobulins are separated, we can move to the next step.
* The second step is based on the technique used. Immunofixation requires electrophoresis to migrate serum proteins in replicate. Then, specific anti-immunoglobulin antisera are used to treat each replicate. For this, the antisera are not placed in a channel, as in electrophoresis, but they are added individually to each migration lane. The presence of a monoclonal immunoglobulin results in the appearance of a narrow band after staining complex precipitates. For example, in the case of an IgG lambda, there will be a narrow band, both on the track on which was deposited anti-G and on which has been deposited with the anti-lambda.
Merits
Immunofixation tends to replaceprotein electrophoresis
Protein electrophoresis is a method for analysing the proteins in a fluid or an extract. The electrophoresis may be performed with a small volume of sample in a number of alternative ways with or without a supporting medium, namely agarose or poly ...
because :
* it is faster (results within three hours);
* it is somewhat more sensitive. Immunofixation may reveal an immunoglobulin missed out by protein electrophoresis, especially at low concentrations (less than 1 gram/litre);
* it can be partially automated and can be used in more laboratories;
* it is more easily read and interpreted.
Demerits
Immunofixation is however only sensitive to immunoglobulins and is more expensive than protein electrophoresis.See also
*Immunoelectrophoresis
Immunoelectrophoresis is a general name for a number of biochemical methods for separation and characterization of proteins based on electrophoresis and reaction with antibodies. All variants of immunoelectrophoresis require immunoglobulins, als ...
References
Sources
* :fr:ImmunofixationExternal links
* - "Immunofixation - serum" * - "Immunofixation - urine" {{Immune_proteins Glycoproteins Immunologic tests