In Vitro Allergy Diagnostics
المؤلف:
Marcello Ciaccio
المصدر:
Clinical and Laboratory Medicine Textbook 2021
الجزء والصفحة:
p504-505
2025-12-13
48
The use of in vitro tests by various methods introduced in recent decades has undoubtedly led to significant progress in diagnosing IgE-mediated allergic syndromes.
Among the advantages of these tests, globally considered, we should point out above all the remarkably high sensitivity and specificity, the good reproducibility, the absolute harm lessness for the patient, and the possibility of being researched even while taking antihistamine therapy; the disadvantages are summarized, in practice, in the relatively high cost.
Detection of Total IgE
The determination of total IgE was introduced many years ago with the PRIST (Paper RadioImmunoSorbent Test) and is currently performed by various automated methods.
The serum concentration of total IgE varies, in non-atopic adults, from 10 to 200 IU/mL; in infants, this concentration is only a few IU/mL, and it progressively increases and reaches adult levels around the tenth year of life.
Total IgE is generally high in allergic syndromes due to IgE-mediated immunoreactions. Notably, the highest values of IgE are found in allergic diseases, but the finding of “nor mal” values of total serum IgE does not exclude the diagnosis of allergic disease. Indeed, many allergic patients have total IgE levels within the normal range.
It should also be mentioned that total IgE is found with altered values in various pathological conditions that are certainly not allergic and physiological or paraphysiological conditions (e.g., in smokers).
Therefore, it can be concluded that the determination of total IgE alone is of little clinical significance in allergic diseases.
Detection of Specific IgE
Since 1967, the year in which the first specific IgE assay was performed by Wide and collaborators, with the birth of the RAST (RadioAllergoSorbent Test), much progress has been made in the field of laboratory allergology diagnostics, through various stages consisting of the development of technologies based on solid phases with high binding capacity; use of enzymatic tracers linked to monoclonal IgE anti bodies with calibration curves made with standards of known titre in quantitative units kUa/L and produced according to an international reference standard (now 3° IS WHO 11/234, formerly 2° IRP WHO 75/502 for total IgE); the introduction of diagnostic systems for the determination of specific IgE with high analytical sensitivity in total automation (third generation test).
Over the years, RAST has been followed by numerous other methods, which have replaced radioactive markers, and the techniques developed have been numerous (ELISA [Enzyme Linked ImmunoSorbent Assay], agglutination, precipitation). These in vitro methods differed significantly in terms of the type of detection system (colorimetric, fluorimetric, and chemiluminescence), the antiserum (mono or polyclonal, single or mixed), the support for carrying the allergen (cellulose polymers, polystyrene spheres, etc.), and the development in a solid or liquid phase, resulting in significant inhomogeneity among the different systems available in the laboratory setting. Since not all systems are the same, and not all are equally valid, the laboratory must guarantee the reproducibility of the data (system with a coefficient of variation < 15% – NCCLS, 2004) and is certified by a national and/or international quality control (QC) program. Currently, the most used methods for detecting specific IgE are Cap FEIA and 3gAllergy.
Cap FEIA on Immunocap
It uses an immunofluorenzyme method and is based on a solid phase (cellulose polymer) covalently conjugated with the allergen, which reacts with the serum under examination so that the specific IgE, if present, binds to the allergen. After washing to remove unbound IgE, i.e., not specific for the allergen, labeled anti-IgE antibodies are added. Thus, a “sandwich” complex consists of the solid phase with allergen + patient-specific IgE + labeled anti-IgE. Using the fluorimetric detection method, the amount of marker present in the complex is measured, which will be directly proportional to the amount of specific IgE present in the sample under examination.
3gAllergy on Immulite 2000
It uses an enzyme-enhanced chemiluminescent method and is based on the use of a liquid phase. Patient serum- containing specific IgE and liquid allergen are incubated in the presence of antiligand coated beads. The liquid allergen, which acts as a ligand, binds to the specific IgE in the serum and the antiligand- coated beads. The ligand-specific IgE is detected by an anti-IgE antibody conjugated to an enzyme that catalyzes the chemiluminescent reaction in the presence of its specific substrate. Also, in this method, the concentrations obtained are proportional to the amount of specific IgE present in the patient’s serum under examination.
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