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Immunological Tests for Diagnosis of Disease and Identification of Molecules
Published in Julius P. Kreier, Infection, Resistance, and Immunity, 2022
The enzyme-linked immunosorbent assay (ELISA) technique is a highly versatile, sensitive, and quantitative procedure that requires little equipment and for which reagents are readily available. It was first introduced in 1971 by Engvall and Perlmann. The term enzyme immunoassay (EIA) is an alternative name for this type of test and is used for the many variants of the assay. Enzymes such as alkaline phosphatase or peroxidase can be linked to antibody without destroying either the antibody′s specificity or the enzyme′s activity. The enzyme acts as a label which makes detection of the antibody possible. Both monoclonal and polyclonal antibodies can be used in this type of test. Enzyme labels are cheaper, simpler to measure, and far more stable than radioactive labels. For these reasons, ELISA or EIA assays have in many cases replaced RIA and have done so while maintaining sensitivity and specificity. Many of the assays for hepatitis A and B in use today are based on ELISA. In addition to the use of enzymes in immunoassays, nonenzy-matic markers can also be used if they can be conjugated to the antibody (e.g., colloidal gold with silver enhancement, and biotination with avidine).
Diagnosis of COVID-19 Using Clinical Examination, Immunoassays, and Molecular Diagnostic Techniques
Published in Srijan Goswami, Chiranjeeb Dey, COVID-19 and SARS-CoV-2, 2022
Srijan Goswami, Ushmita Gupta Bakshi
This serological method takes a much longer time as compared with the rapid test and is useful in gathering epidemiological data, as well as helping to develop treatments. ELISA stands for enzyme-linked immunosorbent assay. In this process, a small amount of serum sample is collected and put into the microtiter well, then IgG from recovered individuals (primary antibody) is added to it. The idea is that the primary antibody will bind to the spike protein antigen of SARS-CoV-2. After that, a secondary antibody tagged with the specific enzyme is added to the reaction mixture. The enzyme-linked secondary antibody binds to the primary antibody. A chromogenic chemical is added to the reaction mixture that acts as the substrate for the enzyme-linked secondary antibody. This enzyme substrate reaction generates a color that is sensed and analyzed by the ELISA reader. The generation of color means a positive test and the intensity of the color is directly proportional to the concentration of antigens. So these are the serology tests that are combined, in addition to RT-PCR (CDC, 2020; Center for Devices and Radiological Health, 2021; Uwamino et al., 2021; Kang et al., 2021).
Hypersensitivity
Published in Peter Kam, Ian Power, Michael J. Cousins, Philip J. Siddal, Principles of Physiology for the Anaesthetist, 2020
Peter Kam, Ian Power, Michael J. Cousins, Philip J. Siddal
Immunodiffusion is used for the qualitative and quantitative analysis of antigens and antibodies in serum or body fluids. The method depends on a precipitation reaction of an insoluble antigen–antibody complex from soluble antigen and antibodies. The enzyme-linked immunosorbent assay (ELISA) is an immunoassay using enzyme-linked antibody or antigen (Figure 58.8). Either the antigen or the antibody can be linked to an enzyme (e.g. horseradish peroxidase or alkaline phosphatase). In the standard indirect ELISA, an antigen is adsorbed onto a polystyrene plate (solid phase). The antibody to be detected binds to this component, and a second enzyme-labelled antibody is then added. The substrate of the enzyme is added, producing a coloured reaction product that can be measured spectrophotometrically.
Antibody tests for COVID-19
Published in Baylor University Medical Center Proceedings, 2021
Jonathan Kopel, Hemant Goyal, Abhilash Perisetti
The antibody tests are a rapid method for detecting antigens (the spike, membrane, or nucleocapsid proteins) or antibodies for COVID-19 using lateral flow assays.3 As shown in Table 1, there are four major types of antibody tests: rapid diagnostic tests (RDT), enzyme-linked immunosorbent assays (ELISA), neutralization assays, and chemiluminescent immunoassays.12 Current World Health Organization guidelines recommend obtaining a blood sample during the first week of illness and then 3 to 4 weeks later to measure SARS-CoV-2 antibodies.13 Within 5 days of infection, the IgM positive rate increased from 50% to 81%, whereas the IgG positive rate increased from 81% to 100% in COVID-19 patients.13 Unlike the nasopharyngeal RT-PCR tests, the antibody tests allow for better collection of epidemiological data, determination of the immune status of asymptomatic individuals, and screening of previous exposure.3 Currently, there is no accepted standard antibody test by which to compare the sensitivity or specificity for the SARS-CoV-2.1 The Food and Drug Administration has required clinics and industries to estimate the analytic performance of any antibody test using a material containing SARS-CoV-2 at sufficient concentration to be detected.
Highly sensitive detection of antibody nonspecific interactions using flow cytometry
Published in mAbs, 2021
Emily K. Makowski, Lina Wu, Alec A. Desai, Peter M. Tessier
There are a number of reported methods for assessing antibody polyspecificity.16–21 The most common ones are enzyme-linked immunosorbent assays (ELISAs), which evaluate binding of antibodies to diverse types of immobilized biomolecules such as proteins, lipids, sugars, DNA, and/or virus particles.16,17 The advantages of ELISAs are their simplicity and ability to evaluate antibody polyspecificity using diverse types of immobilized reagents without the need to label the reagents and detect their binding in solution. However, compared to the detection of highly specific affinity interactions, the sensitivity of ELISAs to detect nonspecific interactions is much weaker, which limits their widespread utility for sensitively evaluating antibody polyspecificity. Several other methods have been reported that adapt traditional methods for measuring affinity protein interactions to evaluating antibody nonspecific interactions, including bio-layer interferometry, surface plasmon resonance, and affinity chromatography (in the form of cross-interaction chromatography).18–21
Three-in-one procedure for failed spinal surgery improved pain, disability scores and serum inflammatory milieu: Three-years follow-up
Published in Egyptian Journal of Anaesthesia, 2021
Serum levels of estimated markers were measured using enzyme-linked immunosorbent assay (ELISA) kits according to the manufacturer’s instructions and were read using a 96 well microplate ELISA reader (Dynatech. MR 7000). Human TNF-α was measured with the enzyme linked immunoassay (ELISA) kit (catalogue no. ab179886, abcam Inc., Cambridge, USA) by quantitative sandwich enzyme immunoassay technique. [27]Human IL-1β with the enzyme linked immunoassay (ELISA) kit (catalogue no. ab46052, abcam Inc., Cambridge, USA) by quantitative sandwich enzyme immunoassay technique. [28]Human IL-6 with the enzyme linked immunoassay (ELISA) kit (catalogue no. ab46042, abcam Inc., Cambridge, USA) by quantitative sandwich enzyme immunoassay technique. [29]Human IL-10 was measured with the enzyme linked immunoassay (ELISA) kit (catalogue no. ab215089, abcam Inc., San Francisco, USA) by quantitative sandwich enzyme immunoassay technique. [30]