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Overview of Cell Culture Processes
Published in Wei-Shou Hu, Cell Culture Bioprocess Engineering, 2020
Antibodies are among the most abundant proteins in blood circulation. They are highly soluble and can be secreted by B cells at high levels. Portions of the antibody molecule, namely the Fc region, are major components of many therapeutic fusion proteins. In those fusion proteins, the functional domain (or fragment) of a protein is joined to the carrier domain through a linker segment (Figure 1.2). The Fc fragment provides many of the properties of an antibody. A prominent example is the fusion molecule of the Fc fragment of IgG and the tumor necrosis factor α (TNFα) binding fragment of the TNFα receptor (TNFRα). The molecule binds to TNFα and suppresses its inflammatory effect. Such non-natural proteins are increasingly being explored as medicine. Bispecific antibodies (BsAbs), as the name implies, use antigen-binding sites from two different antibody molecules to simultaneously target two components of the cellular pathways and thereby improve clinical efficacy. Bispecific T-cell engagers (BiTEs) are a special class of bispecific antibodies which engage the T cells of the host’s immune system in order to treat cancers. Increasingly, antibodies are being derivatized so as to contain drugs. These antibody-drug conjugates (ADCs) deliver cytotoxic agents specifically to diseased cells through their recognition of a particular antigen on the cell surface. Many biologics in the pipeline include antibody-drug conjugates, bispecific antibodies, and bi- or tri-specific immune cell engagers.
GMR Spin-Valve Biosensors
Published in Evgeny Y. Tsymbal, Igor Žutić, Spintronics Handbook: Spin Transport and Magnetism, Second Edition, 2019
Jung-Rok Lee, Richard S. Gaster, Drew A. Hall, Shan X. Wang
The overall structure of IgG antibodies is remarkably similar, whether it is reactive to a cancer tumor marker or an infectious pathogen (Figure 15.1). Every IgG antibody contains two long heavy chains and two short light chains held together via inter-chain disulfide bonds and electrostatic interactions. Both the light chains and the heavy chains can be divided into two distinct regions: a constant region at the base of each chain and a variable region at the tip. The constant region at the base of the heavy chains is typically referred to as the Fc region (which will be mentioned later). At the tip of each chain is the highly variable region known as the Fab region, which contains the antigen-binding sites. This highly variable region is where the specificity of the protein–antibody interaction takes place and the lock-and-key interface occurs. By choosing an antibody that reacts with a specific biomarker of interest, one can selectively capture that biomarker of choice over a sensor surface to facilitate protein detection.
Molecular Analysis in Mechanobiology
Published in Jiro Nagatomi, Eno Essien Ebong, Mechanobiology Handbook, 2018
Antibodies are proteins expressed by B lymphocytes that contribute to specific immune responses by binding to foreign (nonself) molecules termed antigens. The basic antibody subunit is a Y-shaped structure that is composed of two identical copies of heavy and light chains connected by disulfide bridges (Figure 3.4). Five different classes of antibodies are expressed (IgM, IgG, IgD, IgA, and IgE) that differ in the number of Y subunits and the type of heavy chain. Each antibody subunit has two antigen-binding domains in the Fab regions that are identical among all the antibodies produced by an individual B cell, but highly variable among the overall population of B cells. This diversity results from recombination of the immunoglobulin genes during B cell maturation and provides each individual with an antibody repertoire capable of recognizing an estimated >109 different antigens [66]. Each antibody subunit also has an Fc region that is constant among all the antibodies of a particular class and provides binding sites for complement proteins and macrophages. The antigen-binding domains of each antibody recognize a relatively small region of the antigen (typically 6–10 amino acids) termed an epitope. Therefore, a typical antigen such as a protein will contain multiple epitopes, each of which may be recognized by different specific antibodies. Antibody–antigen binding is mediated by multiple, cooperative non-covalent interactions, providing very high specificity that has made antibodies the central tool for studying protein expression.
Cytoplasmic and periplasmic expression of recombinant shark VNAR antibody in Escherichia coli
Published in Preparative Biochemistry and Biotechnology, 2019
Herng C. Leow, Katja Fischer, Yee C. Leow, Katleen Braet, Qin Cheng, James McCarthy
To increase the binding specificity of VNAR towards their target antigens, protein engineering can be deployed to correct erroneous protein from eukaryote proteins in the bacterial expression system.[22] For instances, error-prone PCR and random mutagenesis of the amino acid residues in CDR codon have been used to improve the binding ability and protein functionality in several antibodies, including scFv[80] and camelids VHH.[81] Alternatively, optimization of gene expression system by using different promoters, signal peptides or co-expression of foldase are the other methods used to enhance the solubility and folding capability of the proteins strictly depending on correct disulfide bridges.[20,22] Since lacking appropriate anti-shark antibody the recombinant shark VNAR proteins have been tagged with human constant kappa (HuCκ)[16] or human Fc-region[82] at the C-terminal and expressed in E. coli or mammalian expression system, respectively. These fusion proteins can be detected by commercial available secondary antibody rather than raising anti-shark hybridomas.