The different areas of immunoassays
Whether it is determining a Vitamin-D deficit or the presence of HIV in clinical specimens; there are immunoassays to measure these specific biomarkers. This blog provides an overview of the different immunoassays present in the current market.
Immunoassays and the different options
When discussing immunoassays and the different options in that area, it is sensible to return to the heart of the matter: the way an immunoassay functions. Immunoassays developed by us verify the presence of a specific marker in clinical specimen by using antibodies and antigens. By setting up the right composition a reaction is triggered. In short: any reaction indicates the presence of a specific substance. In case of no reaction, the marker for which the test is created for is not identifiable in the specimen.
A biomarker can be measured in different kind of immunoassays. Although the outcome is similar, the method and technology behind the test may differ greatly. To illustrate this, we discuss several assay models in the following section.
The enzyme-linked immunosorbent assay determines for example the presence of antibodies to an infectious disease. EIA (Enzyme Immunoassay), also known as ELISA (Enzyme Linked Immune Sorbent Assay), detects and measures antibodies in human samples as a reaction to specific antigens. ELISA-tests are also used to test the concentration of, for example, Vitamin D, insulin or hepatitis antigens in samples.
A basic ELISA-test is executed by coating antigens or antibodies to a surface. Next, a sample possibly containing matching antibodies is applied to the surface. Only antibodies present to a specific marker bind to the antigens. Actual binding depends on the presence or absence of these antibodies. All antibodies that do not bind are discarded. What is left is the pure antibodies-antigens connection of interest.
Before starting an ELISA-test the detecting or secondary antibodies are linked to enzymes. The final step in ELISAs consists of adding a substrate that reacts with the enzyme. This produces color formation. The more bindings are formed, the more color is released. The more color, the more a specific substance is present in the sample.
Fluorescence and Chemiluminescence Immunoassays
Besides immunoassays based on color intensity to detect antibodies in specimen, detection can also be done by fluorescent tracers and light-generating molecules. These are called Fluorescent Immunoassay (FIA) and Chemiluminescence Immunoassay (CLIA). The basis of these immunoassays is a variant of ELISA. A substrate is used that does not generate color but generates light.
FIA and CLIA are more sensitive than ELISA and are used to determine low concentrations of antibodies or antigens (HIV, Troponin I, TBI).
A multiplex micro array is a proper option to detect multiple substances. A micro array is a collection of microscopically small spots. These spots are usually deployed in multiplex-assays to do a parallel search for multiple substances, e.g. allergens, infectious diseases and auto-immune diseases. For a client requesting immunoassay development services this form of testing is mainly chosen when it is important to detect several biomarkers simultaneously in blood samples. Read-out can be either colorimetric or fluorescent.
Point of care tests
Point of care tests (PoC) are commonly known as ‘near-patient tests’. This makes sense when you understand that these immunoassays are easily brought to the patient’s location. This makes it possible to quicken patient care by receiving results in a short period of time at the place where necessary. The biggest advantage is to make a well-founded decision in the treatment of the patient. Blood glucose tests, screening for drugs, cholesterol screening and screening for food pathogens and infectious diseases are common examples of PoC tests.
There are different types of PoC-immunoassays. A quantitative PoC test determines the concentration in which a certain substance is present in the human body. A qualitative PoC test determines merely the presence or absence of a certain substance, e.g. a pregnancy test. The tests are categorized as A. elementary tests with a low chance of error measurement or B. more complex tests. The last category is the most commonly used and these tests are therefore always executed by professionals.
Turbidimetry is a method to determine the number of cells in a solution, by measuring the turbidity. This turbidimetric approach is a quick one-step method to follow the complex formed by antigen-antibody reactions. Turbidimetric-based immunoassays are less sensitive than the enzyme-based methods.
Turbidimetry-based immunoassays are applied to detect the presence of, for example, calprotectin in blood.
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If you wish to develop an immunoassay we can:
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- advise you about the different types to reach the best result
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