Hook effect

Last updated
Illustration of hook effect adapted from Schiettecatte et al. Hook effect.png
Illustration of hook effect adapted from Schiettecatte et al.

The hook effect refers to the prozone phenomenon, also known as antibody excess or the Postzone phenomenon, also known as antigen excess. It is an immunologic phenomenon whereby the effectiveness of antibodies to form immune complexes can be impaired when concentrations of an antibody or an antigen are very high. The formation of immune complexes stops increasing with greater concentrations and then decreases at extremely high concentrations, producing a hook shape on a graph of measurements. An important practical relevance of the phenomenon is as a type of interference that plagues certain immunoassays and nephelometric assays, resulting in false negatives or inaccurately low results. Other common forms of interference include antibody interference, cross-reactivity and signal interference. The phenomenon is caused by very high concentrations of a particular analyte or antibody and is most prevalent in one-step (sandwich) immunoassays. [2] [3]

Contents

Mechanism and in vitro importance

Prozone - excess antibodies

Illustration of the effects of excess antigen and blocking antibodies on immunoassays. Illustration of the effects of excess antigen and blocking antibodies on immunoassays.png
Illustration of the effects of excess antigen and blocking antibodies on immunoassays.

In an agglutination test, a person's serum (which contains antibodies) is added to a test tube, which contains a particular antigen. If the antibodies interact with the antigen to form immune complexes, called agglutination, then the test is interpreted as positive. However, if too many antibodies are present that can bind to the antigen, then the antigenic sites are coated by antibodies, and few or no antibodies directed toward the pathogen are able to bind more than one antigenic particle. [4] Since the antibodies do not bridge between antigens, no agglutination occurs. Because no agglutination occurs, the test is interpreted as negative. In this case, the result is a false negative. The range of relatively high antibody concentrations within which no reaction occurs is called the prozone. [5]

Postzone - excess antigens

The effect can also occur because of antigen excess, when both the capture and detection antibodies become saturated by the high analyte concentration. In this case, no sandwich can be formed by the capturing antibody, the antigen and the detection antibody. In this case, free antigen is in competition with captured antigen for detection antibody binding. [6] Sequential addition of antigen and antibody, paired with stringent washing, can prevent the effect, as can increasing the relative concentration of antibody to antigen, thereby mediating the effect.[ citation needed ]

Simple illustration of the effects of excess antigen and dosage response curve. Typical Hook effect illustration.png
Simple illustration of the effects of excess antigen and dosage response curve.

Examples include high levels of syphilis antibodies in HIV patients or high levels of cryptococcal antigen leading to false negative tests in undiluted samples. [7] [8] This phenomenon is also seen in serological tests for Brucellosis.[ citation needed ] It may be seen in precipitation reactions. The antibody that fails to react is known as the blocking antibody and prevents the precipitating antibody from binding to the antigens. Thus the proper precipitation reaction does not take place. However, when the serum is diluted, the blocking antibody is as well and its concentration decreases enough for the proper precipitation reaction to occur. [9]

In vivo observations

Lewis Thomas described in his memoir a physiologic experiment of 1941 in which he observed the prozone effect in vivo: immunity in rabbits to meningococcus, which was robust, unexpectedly decreased when immunization was used to induce a heightened antibody response. [10] In other words, getting the rabbits' bodies to produce more antibodies against this bacterium had the counterproductive effect of decreasing their immunity to it. From the viewpoint of an overly simplistic notion of the antibody/antigen relationship, this seems paradoxical, although it is clearly logical from a viewpoint duly informed by present-day molecular biology. Thomas was interested in pursuing this physiologic research further, and remained so for decades afterward, but his career took him in other directions and he was not aware of anyone having pursued it by the time of his memoir. [10] One kind of relevance that he hypothesized for this in vivo blocking antibody concept was as a driver of human susceptibility to certain infectious diseases. [10] In the decades since, the concept has also been found to have clinical relevance in allergen immunotherapy, where blocking antibodies can interfere with other antibodies involved in hypersensitivity and thus improve allergy treatment. [11]

See also

Related Research Articles

<span class="mw-page-title-main">ELISA</span> Method to detect an antigen using an antibody and enzyme

The enzyme-linked immunosorbent assay (ELISA) is a commonly used analytical biochemistry assay, first described by Eva Engvall and Peter Perlmann in 1971. The assay uses a solid-phase type of enzyme immunoassay (EIA) to detect the presence of a ligand in a liquid sample using antibodies directed against the protein to be measured. ELISA has been used as a diagnostic tool in medicine, plant pathology, and biotechnology, as well as a quality control check in various industries.

An assay is an investigative (analytic) procedure in laboratory medicine, mining, pharmacology, environmental biology and molecular biology for qualitatively assessing or quantitatively measuring the presence, amount, or functional activity of a target entity. The measured entity is often called the analyte, the measurand, or the target of the assay. The analyte can be a drug, biochemical substance, chemical element or compound, or cell in an organism or organic sample. An assay usually aims to measure an analyte's intensive property and express it in the relevant measurement unit.

<span class="mw-page-title-main">Seroconversion</span> Development of specific antibodies in the blood serum as a result of infection or immunization

In immunology, seroconversion is the development of specific antibodies in the blood serum as a result of infection or immunization, including vaccination. During infection or immunization, antigens enter the blood, and the immune system begins to produce antibodies in response. Before seroconversion, the antigen itself may or may not be detectable, but the antibody is absent. During seroconversion, the antibody is present but not yet detectable. After seroconversion, the antibody is detectable by standard techniques and remains detectable unless the individual seroreverts. Seroreversion, or loss of antibody detectability, can occur due to weakening of the immune system or waning antibody concentration over time. Seroconversion refers the production of specific antibodies against specific antigens, meaning that a single infection could cause multiple waves of seroconversion against different antigens. Similarly, a single antigen could cause multiple waves of seroconversion with different classes of antibodies. For example, most antigens prompt seroconversion for the IgM class of antibodies first, and subsequently the IgG class.

Serology is the scientific study of serum and other body fluids. In practice, the term usually refers to the diagnostic identification of antibodies in the serum. Such antibodies are typically formed in response to an infection, against other foreign proteins, or to one's own proteins. In either case, the procedure is simple.

<span class="mw-page-title-main">Radioimmunoassay</span> Immunoassay that uses radiolabeled molecules

A radioimmunoassay (RIA) is an immunoassay that uses radiolabeled molecules in a stepwise formation of immune complexes. A RIA is a very sensitive in vitro assay technique used to measure concentrations of substances, usually measuring antigen concentrations by use of antibodies.

<span class="mw-page-title-main">Immunoassay</span> Biochemical test for a protein or other molecule using an antibody

An immunoassay (IA) is a biochemical test that measures the presence or concentration of a macromolecule or a small molecule in a solution through the use of an antibody (usually) or an antigen (sometimes). The molecule detected by the immunoassay is often referred to as an "analyte" and is in many cases a protein, although it may be other kinds of molecules, of different sizes and types, as long as the proper antibodies that have the required properties for the assay are developed. Analytes in biological liquids such as serum or urine are frequently measured using immunoassays for medical and research purposes.

The direct and indirect Coombs tests, also known as antiglobulin test (AGT), are blood tests used in immunohematology. The direct Coombs test detects antibodies that are stuck to the surface of the red blood cells. Since these antibodies sometimes destroy red blood cells they can cause anemia; this test can help clarify the condition. The indirect Coombs test detects antibodies that are floating freely in the blood. These antibodies could act against certain red blood cells; the test can be carried out to diagnose reactions to a blood transfusion.

<span class="mw-page-title-main">Serotype</span> Distinct variation within a species of bacteria or virus or among immune cells

A serotype or serovar is a distinct variation within a species of bacteria or virus or among immune cells of different individuals. These microorganisms, viruses, or cells are classified together based on their surface antigens, allowing the epidemiologic classification of organisms to a level below the species. A group of serovars with common antigens is called a serogroup or sometimes serocomplex.

Heterophile antibodies are antibodies induced by external antigens.

Cross-reactivity, in a general sense, is the reactivity of an observed agent which initiates reactions outside the main reaction expected. This has implications for any kind of test or assay, including diagnostic tests in medicine, and can be a cause of false positives. In immunology, the definition of cross-reactivity refers specifically to the reaction of the immune system to antigens. There can be cross-reactivity between the immune system and the antigens of two different pathogens, or between one pathogen and proteins on non-pathogens, which in some cases can be the cause of allergies.

<span class="mw-page-title-main">Ouchterlony double immunodiffusion</span>

Ouchterlony double immunodiffusion is an immunological technique used in the detection, identification and quantification of antibodies and antigens, such as immunoglobulins and extractable nuclear antigens. The technique is named after Örjan Ouchterlony, the Swedish physician who developed the test in 1948 to evaluate the production diphtheria toxins from isolated bacteria.

A blocking antibody is an antibody that does not have a reaction when combined with an antigen, but prevents other antibodies from combining with that antigen. This function of blocking antibodies has had a variety of clinical and experimental uses.

A precipitin is an antibody which can precipitate out of a solution upon antigen binding.

A latex fixation test, also called a latex agglutination assay or test, is an assay used clinically in the identification and typing of many important microorganisms. These tests use the patient's antigen-antibody immune response. This response occurs when the body detects a pathogen and forms an antibody specific to an identified antigen present on the surface of the pathogen.

Human anti-mouse antibody or human anti-murine antibody (HAMA) is an antibody found in humans which reacts to immunoglobins found in mice.

<span class="mw-page-title-main">Heterophile antibody test</span> Diagnosistic test for infectious mononucleosis

The mononuclear spot test or monospot test, a form of the heterophile antibody test, is a rapid test for infectious mononucleosis due to Epstein–Barr virus (EBV). It is an improvement on the Paul–Bunnell test. The test is specific for heterophile antibodies produced by the human immune system in response to EBV infection. Commercially available test kits are 70–92% sensitive and 96–100% specific, with a lower sensitivity in the first two weeks after clinical symptoms begin.

A nontreponemal test (NTT) is a blood test for diagnosis of infection with syphilis. Nontreponemal tests are an indirect method in that they detect biomarkers that are released during cellular damage that occurs from the syphilis spirochete. In contrast, treponemal tests look for antibodies that are a direct result of the infection thus, anti-treponeme IgG, IgM and to a lesser degree IgA. Nontreponemal tests are screening tests, very rapid and relatively simple, but need to be confirmed by treponemal tests. Centers for Disease Control and Prevention (CDC)-approved standard tests include the VDRL test, the rapid plasma reagin (RPR) test, the unheated serum reagin (USR) test, and the toluidine red unheated serum test (TRUST). These have mostly replaced the first nontreponemal test, the Wassermann test.

Antigen-antibody interaction, or antigen-antibody reaction, is a specific chemical interaction between antibodies produced by B cells of the white blood cells and antigens during immune reaction. The antigens and antibodies combine by a process called agglutination. It is the fundamental reaction in the body by which the body is protected from complex foreign molecules, such as pathogens and their chemical toxins. In the blood, the antigens are specifically and with high affinity bound by antibodies to form an antigen-antibody complex. The immune complex is then transported to cellular systems where it can be destroyed or deactivated.

<span class="mw-page-title-main">Fluorescence polarization immunoassay</span> Class of invitro biochemical test

Fluorescence polarization immunoassay (FPIA) is a class of in vitro biochemical test used for rapid detection of antibody or antigen in sample. FPIA is a competitive homogenous assay, that consists of a simple prepare and read method, without the requirement of separation or washing steps.

<span class="mw-page-title-main">Blood compatibility testing</span> Testing to identify incompatibilities between blood types

Blood compatibility testing is conducted in a medical laboratory to identify potential incompatibilities between blood group systems in blood transfusion. It is also used to diagnose and prevent some complications of pregnancy that can occur when the baby has a different blood group from the mother. Blood compatibility testing includes blood typing, which detects the antigens on red blood cells that determine a person's blood type; testing for unexpected antibodies against blood group antigens ; and, in the case of blood transfusions, mixing the recipient's plasma with the donor's red blood cells to detect incompatibilities (crossmatching). Routine blood typing involves determining the ABO and RhD type, and involves both identification of ABO antigens on red blood cells and identification of ABO antibodies in the plasma. Other blood group antigens may be tested for in specific clinical situations.

References

  1. ↑ Schiettecatte, Johan; Anckaert, Ellen; Smitz, Johan (2012-03-23). "Interferences in Immunoassays". Advances in Immunoassay Technology. InTech. doi:10.5772/35797. ISBN   978-953-51-0440-7.
  2. ↑ Greenberg, G R; Jeejeebhoy, K N (1989-03-01). "Reply". Gut. 30 (3): 422–423. doi:10.1136/gut.30.3.422-a. ISSN   0017-5749. PMC   1378475 .
  3. ↑ Jacobs, Joannes F. M.; van der Molen, Renate G.; Bossuyt, Xavier; Damoiseaux, Jan (2015-02-01). "Antigen excess in modern immunoassays: To anticipate on the unexpected". Autoimmunity Reviews. 14 (2): 160–167. doi:10.1016/j.autrev.2014.10.018. ISSN   1568-9972. PMID   25461469.
  4. ↑ Miller, James (April 2004). "Interference in immunoassays: avoiding erroneous results" (PDF). CLI Online. CLI. Retrieved 3 February 2016.
  5. ↑ Gillet, Philippe; Mori, Marcella; Esbroeck, Marjan Van; Ende, Jef; Jacobs, Jan (2009-11-30). "Assessment of the prozone effect in malaria rapid diagnostic tests". Malaria Journal. 8 (1): 271. doi: 10.1186/1475-2875-8-271 . PMC   2789093 . PMID   19948018.
  6. ↑ Tate, Jill; Ward, Greg (2004-05-01). "Interferences in Immunoassay". The Clinical Biochemist Reviews. 25 (2): 105–120. ISSN   0159-8090. PMC   1904417 . PMID   18458713.
  7. ↑ Jurado RL, Campbell J, Martin PD (November 1993). "Prozone phenomenon in secondary syphilis. Has its time arrived?". Arch. Intern. Med. 153 (21): 2496–8. doi:10.1001/archinte.153.21.2496. PMID   7832818.
  8. ↑ Stamm AM, Polt SS (September 1980). "False-negative cryptococcal antigen test". JAMA. 244 (12): 1359. doi:10.1001/jama.244.12.1359. PMID   6997519.
  9. ↑ Cole, L. A.; Shahabi, S.; Butler, S. A.; Mitchell, H.; Newlands, E. S.; Behrman, H. R.; Verrill, H. L. (2001-02-01). "Utility of commonly used commercial human chorionic gonadotropin immunoassays in the diagnosis and management of trophoblastic diseases". Clinical Chemistry. 47 (2): 308–315. doi: 10.1093/clinchem/47.2.308 . ISSN   0009-9147. PMID   11159780.
  10. 1 2 3 Thomas, Lewis (1983), The Youngest Science: Notes of a Medicine-Watcher
  11. ↑ Strait, RT; Morris, SC; Finkelman, FD (2006), "IgG-blocking antibodies inhibit IgE-mediated anaphylaxis in vivo through both antigen interception and Fc gamma RIIb cross-linking", J Clin Invest, 116 (3): 833–841, doi:10.1172/JCI25575, PMC   1378186 , PMID   16498503
This page is based on this Wikipedia article
Text is available under the CC BY-SA 4.0 license; additional terms may apply.
Images, videos and audio are available under their respective licenses.