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Lateral Flow Assays
Published in Sibel A. Ozkan, Bengi Uslu, Mustafa Kemal Sezgintürk, Biosensors, 2023
Kamil Żukowski, Marcin Drozd, Robert Ziółkowski, Mariusz Pietrzak, Katarzyna Tokarska, Adam Nowiński, Elżbieta Malinowska
However, a number of nanoparticle-based labels does not show intrinsic affinity to bioreceptors. In their case, the immobilization involves functional groups present on the surface, which are active in the coupling reactions. It can be carried out, among others, by amino coupling via carbodiimide chemistry or coupling via reduced sulfhydryl groups of proteins or DNA receptors with terminal amino- or thiol link (2, 71). An attractive alternative is the use of indirect methods, such as binding of antibodies through surface-coated A/G protein or capturing of DNA-biotin conjugates by streptavidin. In the first approach, bacterial proteins are harnessed to bind Fc antibody fragments. Due to the oriented immobilization, Fab fragments are better exposed and they can more easily interact with antigens. On the other hand, the interaction of streptavidin with biotinylated receptors combines a strong binding with a really small negative effect of immobilization on the bioreceptor affinity. Selected methods of biorecognition elements conjugation with popular LFA labels are listed in Table 10.1.
Antibody-based Radionuclide Imaging
Published in Michael Ljungberg, Handbook of Nuclear Medicine and Molecular Imaging for Physicists, 2022
Steffie M.B. Peters, Erik H. J. G. Aarntzen, Sandra Heskamp
Antibodies are heavy (~150 kDa) and Y-shaped globular plasma proteins, predominantly referred to as immunoglobulins (Ig). They consist of two regions, the antigen-binding fragment (Fab region) and the fragment crystallizable (Fc) region (Figure 18.1). The variable region of the Fab part is responsible for binding to specific antigens, while the Fc region interacts with immune cells and proteins. The Fc region also determines the antibody isotype: IgG, IgA, IgM, IgE, or IgD, each of which differs in function and distribution in the human body. For imaging and therapy, IgG is the most commonly used antibody isotype. Within IgG there are four subclasses: IgG1, IgG2, IgG3, and IgG4, each of which has a distinct role in the immune response. While the subtypes share 90 per cent similarity, small structural differences can lead to substantial changes in their in vivo behaviour. For example, the serum half-life of IgG1, IgG2, and IgG4 is approximately 21 days, while for IgG3 this is only 7 days [20, 21].
Role of Engineered Proteins as Therapeutic Formulations
Published in Peter Grunwald, Pharmaceutical Biocatalysis, 2019
Khushboo Gulati, Krishna Mohan Poluri
Antibodies or immunoglobulins (Ig) are the important members of immune system that neutralize the antigens by recognizing their epitopes. Antigen binding site is located in hypervariable regions or complementarity determining regions (CDRs) that mainly involve first 110 amino acids of heavy and light chains of antibodies. Arrangement of variable regions results in diversity among the antibodies (Owen et al., 2012). Several methods have been developed to engineer antibodies with high specificity and affinity (He and Zhu, 2015). Regions including fragments of antigen binding (Fab) comprising VH, VL, CH1, and CL domains and Fragment of crystallization (Fc region) consisting of hinge, CH2, and CH3 domains are the main targets to engineer novel antibodies. Several strategies are also being employed to increase the in vivo half- life of antibodies. Eculizumab is the first antibody with engineered Fc region that was approved in 2007 (Czajkowsky et al., 2012; Wang et al., 2018). In several cases, monoclonal antibodies are not as effective as therapeutic agents, hence, antibody scaffolds are being used to incorporate the specific activity to develop them as successful therapeutic candidates (Gebauer and Skerra, 2009).
Construction and evaluation of wild and mutant ofatumumab scFvs against the human CD20 antigen
Published in Preparative Biochemistry & Biotechnology, 2023
Reza Maleki, Azam Rahimpour, Masoumeh Rajabibazl
Using whole IgG antibodies could come with some drawbacks as therapeutic anticancer agents, including low blood clearance and high production costs, respectively due to their large size and the need for expression in mammalian cells.[7] There are many techniques developed to eliminate these constraints. For example, recombinant fragments of the antibody such as minibodies (scFv-CH3), Fabs (fragment antigen-binding), and single chain variable fragments (scFv) may diminish the cost of production, increase penetration and relatively, prevent the immunogenicity.[8] As well as this, the E. Coli expression method for antibody generation provides a number of plus points including high rates of expression, affordable cultivation expenses, swift growth, low risk of contamination with viral DNA, non-glycosylation, and simpler purification.[9]