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Immunoassay-Based Detection of Infectious and Parasitic Diseases
Published in Richard O’Kennedy, Caroline Murphy, Immunoassays, 2017
The use polyclonal reagents is compromised by the broad specificities generated, the requirement for numerous laboratory animals and batch-to-batch variation. Monoclonal antibody generation is a route to generation of specific antibodies, which is countered by reduced sensitivity, in comparison to the broad coverage of polyclonal antibodies. Classically, such reagents exhibit low nanomolar affinity due to the concept of the affinity ceiling [75]. These limitations can be addressed by recombinant antibody technologies which have allowed the development of extremely high affinity–specific reagents by a number of display technologies [76]. Coupled with advanced approaches for reformatting, these recombinant antibodies can be of considerable importance to diagnostic applications. The development of recombinant antibody-based diagnostic tests is anticipated to come to the fore in the near future. To address cross-reactivity, a knowledge driven approach based upon detailed target evaluation can minimise, or if required maximise target cross-reactivity. The use of alternative hosts for antibody generation, such as chickens, can also assist in reducing false positives due to complement elements and human anti-mouse antibody (HAMA) crosslinking [77]. Thus the quality of the reagents generated will impact significantly on both the sensitivity and specificity of the designed assay. The sample matrix and its preparation combined with the signal detection methodology applied will have a dramatic effect on the sensitivity and several of the approaches outlined in Table 7.2 show sophisticated methodologies to improve assays for infectious disease.
E. coli from drinking water
Published in Cara Gleeson, Nick Gray, The Coliform Index and Waterborne Disease, 1996
Immunodetection assays may also be used for the detection of coliforms and E. coli. Recent decades have seen a tremendous expansion in the number of such techniques available, including agglutination, Radio Immuno Assays (RIA), Enzyme Linked Immuno-Sorbent Assays (ELISA), Immuno-Fluorescent techniques (IF), Immune electron microscopy-immunogold, Immuno-Enzyme Assays (IEA), Monoclonal Antibodies (MoAbs), counter immunoeletrophoresis and neutralization (Kfir, Du Preez and Genthe, 1993). To date, there is extensive use of these tools in the water field. However, they are largely used for the identification and enumeration of pathogens and are rarely used for the routine analysis of indicator organisms (Kfir, Du Preez and Genthe, 1993). Joret et al. (1989) used monoclonal antibodies directed against outer membrane proteins (OMP-F protein) and those directed against alkaline phosphatase (an enzyme located in the cell periplasmic space) to detect coliforms and E. coli. They found anti-porin MoAbs were unable to distinguish between viable and non-viable cells. The anti-alkaline phosphate MoAbs were very specific and allowed rapid visualization. However, it was felt that sensitivity of the method needed to be determined before the method could be used for routine water analysis. Kfir, Du Preez and Genthe (1993) have reviewed in considerable depth the use of many of the immunodiagnostic tools available for water analysis. They concluded that while these methods are in many ways simpler, more rapid and less labour intensive than conventional methods, there are also considerable drawbacks associated with their use. Immunodetection methods are highly specific, but false positive results are a frequent occurrence. This is due to reactions with non-specific matter or cross-reactivity with a wide range of organisms present in a sample. Many methods do not detect viability. Because of these limitations it is unlikely that immunodetection techniques will be used on a routine basis for water analysis.
Design of an effective piezoelectric microcantilever biosensor for rapid detection of COVID-19
Published in Journal of Medical Engineering & Technology, 2021
Hannaneh Kabir, Mohsen Merati, Mohammad J. Abdekhodaie
When a specific agent reactivity stimulates reactions outside the main supposed reaction, cross-reactivity happens. In immunology, this event can be observed between the immune system and antigens when an antibody conjugates with antigens of two different pathogens [47]. The designed biosensor in this study should possess a good functionality in response to cross-reactivity and must be selective enough to differentiate antigens binding to the same antibody. Co-occurrence of COVID-19 pandemics and influenza viruses (flu) which become widespread in fall and winter, might affect a lot of people and must be considered seriously. Influenza viruses are divided into four types of A, B, C, and D. Influenza A and B are the flu seasons causing seasonal epidemic disease. Among these types, only influenza A leads to a flu pandemic. H1N1 is one of the main subtypes of influenza A spreading between people, routinely [48]. Although COVID-19 and H1N1 are originated from different viruses, they cause respiratory disease with a wide range of analogous symptoms from mild to severe disease and death. As they have similar presentations, discriminating between them is difficult which makes testing inevitable. Since COVID-19 has a high speed of transmission, the patients must be distinguished and isolated rapidly in order to control the rate of spreading. In the following, by assessing the H1N1 experimental data, it will be approved that the designed biosensor has the capability to diagnose appropriately if both of the viruses’ antigens tend to pair the same antibody immobilised to the top surface of the microcantilever. Lee and co-workers introduced 32D6 as a human H1N1 neutralising antibody through an Eptein-Barr virus – immobilised B cell-based technology. According to their experimental data, 0.036 µg/ml of 32D6 was needed to neutralise H1N1. Table 3 also displayed the binding kinetics of 32D6 towards HA of H1N1. As it is shown, the dissociation constant (KD) was reported 3.528* 10−10 M. KD is the ratio of Koff/Kon. Kon means how quickly the antibody binds to the antigen and Koff is used to determine how quickly an antibody dissociates from its target. A low KD value discloses a high speed reaction, while a high KD value relates to a low speed interaction. In this research, Kon and Koff were measured as 2.585*105 M−1S−1 and 9.121*10−5S−1, respectively. The low KD value represents a high affinity interaction in this research [49].