The wide field of antibodies consists of a variety of manifold technical terms, the exact meaning of which does not always become accessible at the first glance. This is why we have collected some of them, adding a brief description of their key aspects.
Antibody-dependent cell-mediated cytotoxicity or antibody-dependent cellular cytotoxicity. It is a mechanism of immune response, which includes the binding of antibodies to surface antigens of a pathogen and subsequent strong activation of natural killer cells towards this pathogen. Low fucose content in the Fc region N-glycans enhances ADCC, see: afucosylated antibody.
Adherent cells are cells that require attachment to a surface or structural network to grow. Adherent cells are usually derived from tissue cells, as opposed to e. g. blood cells that float in plasma.
Antibodies that are specifically designed and produced to lack the carbohydrate fucose in their Fc region N-glycans. They trigger stronger ADCC responses than antibodies with native fucose content. The ProBioGen GlymaxX technology allows manufacturers of antibodies to produce afucosylated antibodies without any deviations from the desired amino acid sequence.
Learn more about our afucosylation service
Antibodies are “Y”-shaped glycoproteins consisting of four amino acid chains: two identical heavy chains and two identical light chains that are linked together. They exhibit very high specificities and affinities towards their respective antigens. They are key players in the adaptive immune system of vertebrates and are involved in numerous pathways that defend against pathogens, e.g. directly by tight binding and covering their surface and indirectly via recruitment and activation of immune cells.
Their role in the human immune system and their molecular specificity led to the development of many therapeutics against cancer and autoimmune diseases as well as crucial diagnostic tools.
Antibody expression is the production of antibody molecules in ribosomes from single amino acid building blocks according to RNA blueprints.
This process has been harnessed by molecular biology and biotechnology to manufacture enhanced and engineered antibodies in cell cultures for therapeutic uses and as diagnostic and research tools.
Antigens are molecular and cellular features that are tightly bound by antibodies’ “antigen binding site”. Common antigens are toxins, pathogens, virus components, bacterial cell walls, but antibodies can be engineered to bind virtually any chemical or biological structure, which then becomes their respective antigen.
The region at the two tips of antibodies “Y” that binds the antigen. They are formed by the variable domains of the Fab fragments (fragment antigen binding). Since the antigen-binding site confers the remarkably high specificity of antibodies, this is the region that varies the most between different antibodies and each contains three hypervariable regions.
Anti-idiotypic antibodies bind the idiotype of another antibody, usually an antibody drug. The idiotype of an antibody is a part of its variable region, hence anti-idiotypic antibodies are rather specific for their target. Depending on the binding geometry, three types of anti-idiotypic antibodies are defined: the antigen-blocking type binds directly in and occupies the target’s antigen binding site, non-blocking type binds laterally and allows antigen binding in the target’s antigen binding site, and complex specific, which binds the target when bound to its antigen.
API is the component of a drug formulation (i. e. tablet, injectable solution) that is pharmaceutically active and leading to therapeutic (and concomitant adverse side effects). It is usually, e. g. mixed with excipients, filled into capsules or dissolved in a suitable solvent to make the active molecule better suited for efficient resorption into the patient’s body.
B cells, or B lymphocytes are crucial constituents of the immune system. In humans they are produced from stem cells in the bone marrow of big hollow bones. They produce antibodies which are either attached to their cellular surface or secreted into their surroundings (tissues or plasma). Various subtypes of B cells are known: from short lived plasmablasts that secrete antibodies in early stages of infection to long lived memory cells that remain after a cured infection and trigger a strong immune response upon reactivation.
Also known as biologics. A class of therapeutics that contains therapeutic medical products that are produced, developed or engineered from biological systems or their biomolecules. The global market size of biotherapeutics is estimated at over 100 billion US dollars and estimated to double until the end of the present decade. Biotherapeutic research stimulates the life sciences, medicine, pharmaceutical sciences and (bio-)process technology, and facilitated the development of numerous first-in-class therapies, having a huge impact on uncounted patients’ lives.
Bispecific antibodies are synthetic antibodies that are highly specific towards two distinct antigens, whereas natural antibodies bind one antigen. They are produced by fusing the genetic blueprints of two distinct antibodies using recombinant and molecular biology technologies. The resulting gene is then expressed in cell culture. Bispecific antibodies are promising candidates for therapeutic uses and as research tools.
Chimeric antibodies are made by humanizing non-human antibodies. Historically, animals were the single available source of antibodies after being exposed to an antigen. The resulting antibodies had traits of the host animals and triggered immune responses when administered to patients. Thus, such antibody drugs need to have human antibody traits while keeping antigen binding fragments of non-human antibodies.
The Chinese Hamster (Cricetulus griseus) is a small rodent native to Mongolia and China. They weigh from 30 g to less than 50 g, with a body length of 8 to 13 cm with a long tail that distinguishes it from other hamsters. They are famous in life sciences because the very popular and widely used CHO cell lines were derived from Chinese Hamster Ovary cells.
A mammalian cell line derived from Chinese Hamster Ovary cells. They are widely used in mammalian cell cultures for recombinant protein expression due to their relative ease in handling and high protein yields. Moreover, as eukaryotic cells they allow the expression of more complex proteins with native post-translational modifications, which are not accessible via E. coli cell cultures. In addition, the use of CHO cells abolishes the widespread need for laboratory animals for antibody production.
CHO cell antibody production refers to the process of using CHO cells to produce large quantities of a specific antibody. This process typically involves genetic engineering of the CHO cells to express the gene encoding the antibody of interest, followed by cultivation of the cells under controlled conditions to promote growth and production of the antibody.
The complement system is an innate branch of the human immune system that is complementary to the adaptive immune system. The complement system does not rely on antibodies or specific cells but consists of various protein precursors, which, when activated by pathogens, lead to inflammatory responses and recruitment of phagocytes to remove intruders or damaged cells.
Cell culture medium is the substrate in which cells are cultured. Depending on the cell type, the culture medium may be a gel (agar medium) or a liquid that provides a suitable environment for the cultured cells. It provides the cells with nutrients (energy and nitrogen sources), vitamins, minerals, salts and keeps osmotic pressure in physiologic ranges. Additionally, it dilutes metabolic waste products. Mammalian cells may require specific gas pressures of oxygen and carbon dioxide plus the presence of hormones to ensure optimal growth conditions.
Enzymes are proteins that accelerate a specific chemical reaction, i. e. they act as biological catalysts. In general, they are relatively large polypeptides, made of one or several chains of amino acids that fold into a specific 3D structure. They are specific to their substrates and catalyzed reaction type. Virtually any process in a cell is catalyzed and therefore regulated by an enzyme. Prominent examples include digestive enzymes that chop food into suitable nutrients, enzymes in the citric acid cycle that provide energy to cells, and enzymes that regulate gene expression.
Epitope denotes the region of an antigen that is recognized and bound by an antibody. The region of an antibody that binds to the epitope is called paratope.
The Fab region of an antibody contains the paratope and consists of part of one heavy and one light chains, specifically one constant region and one variable region. One Fab corresponds to one tip of an antibodies’ “Y”.
Fabs hold promise as therapeutics, e. g. as antidotes for venom.
Fc receptors are found on the surface of several immune cells, e. g. B cells and natural killer cells (NK cells). They are bound and activated by the fragment crystallizable (Fc) region of antibodies. This interaction is the activating mechanism of ADCC, in which NK cells are strongly recruited by Fc regions with artificially low fucose content, leading to fierce activation of the immune system.
Fc-silencing is the minimization of the Fc-dependent recruitment of immune system cells through antibody-antigen complexes. Many promising therapeutic approaches rely on the mere inactivation of a biological process through antibody binding. In the past, these approaches were not feasible due to concomitant activation of the immune system, leading to adverse side effects. With the development of Fc-silencing, these therapies gained tremendous momentum. Truly Fc silent antibodies are accessible through STR technology, developed by mAbsolve, with whom evitria maintains a partnership.
Learn more about our fc-silencing service
Gene expression is the biological process of protein production, involving transcription of genetic blueprints into RNA strands and subsequent synthesis of polypeptides from single amino acids in the ribosome.
Good Manufacturing Practices (GMP) are guidelines which are recommended by specific regulatory agencies, e. g. the US Food and Drug Administration (FDA). These guidelines encompass processes and measures to ensure consistent high quality of products with the end goal of preventing harm to the end customer, e. g. a patient receiving a pharmaceutical product.
Glycoengineered antibodies contain modifications in the glycosylation (N-glycan portion) of the molecular structure. Since the glycosylation pattern plays a key role in the signaling pathways of the immune system, controlling it is crucial in the manufacture of antibodies. Antibodies from prokaryotic cell cultures do not contain glycosylation at all and are inferior to antibodies from mammalian cell cultures. Afucosylated antibodies and Fc-silenced antibodies lie at opposing ends of the glycoengineering spectrum: the first are strong immune system activators, the latter exhibit no activation.
Proteins that are linked to glycans (oligosaccharides, or sugar-chains). Common synonyms: glycosylated proteins, N-glycan containing proteins.
Glycosylation is the term that denotes the linking of glycans (chains of sugar molecules) to another (bio-)molecule. In the context of antibodies, usually the N-glycosylation of e. g. the Fc region is implied.
Heavy chains are components of antibodies, which consist of two heavy chains and two light chains. Each heavy chain spans the whole length from the base to the tip of the “Y” of an antibody and contributes to the antigen binding as well as the signaling mechanisms at the Fc region.
HEK-293 is the acronym for Human Embryonic Kidney cells. HEK cell lines were developed from a single tissue sample by means of treating them with adenovirus. Experiment No. 293 yielded immortalized cells that continued to grow and multiply in suitable cell culture conditions. They are relatively easy to culture and are popular for cell experiments as well as for gene expression. When comparing HEK293 vs. CHO cells, both have unique properties that qualify them as recombinant antibody expression systems.
Hybridoma is a type of immortalized cell that emerges from fusion of a B cell and a myeloma cell and produces antibodies. It was popular in monoclonal antibody production prior to recombinant technology. B cells were harvested from immunized animals and selected for desirable antibody properties. Suitable cells were fused with myeloma cells to generate a cell line that combines the antibody producing properties of the B cell and the immortality of the myeloma cell. The resulting hybridoma cells were injected into animals or grown in culture to continually harvest the secreted antibodies.
The immune system is a highly complex network of cells and proteins that defend the organism against pathogens like bacteria, viruses, parasites and toxins. One branch of the human immune system is adaptive in the sense that once exposed to a pathogen, it is rapidly recognized and a strong immune response is triggered. Both processes are mediated by antibodies, that have the ability to recognize molecular features with high specificity and recruit immune cells to digest.
In vitro denotes processes like experiments or production steps that are performed outside of an organism, traditionally in glass tubes or flasks, hence “vitro” meaning “glass”.
In vivo describes processes that occur in the living organism. Sometimes used in contrast to “in vitro” to imply more natural circumstances of an experiment or a study.
Immunoglobulins (Ig) are a synonymous term for antibodies. There are five isotypes of Igs: IgG, IgA, IgM, IgE and IgD with a specific type of heavy chain. Their distribution among tissues and their role in the immune system is distinct from each other.
Lymphocytes are a subtype of leukocytes and are constituents of the human immune system. Prominent lymphocytes are natural killer cells (NK cells, innate immune system), B cells and T cells, both part of the adaptive immune system.
Mammalian cell lines are derived from mammals and can be grown in cell cultures. They are not as easily grown as simpler cell types, but the expressed antibodies are of much higher quality due to the native glycosylation pattern.
Monoclonal antibodies are antibodies that are virtually identical in their affinity and specificity against their antigen and bind one single epitope. They are generally produced by cells which originate from the same cell, hence share the same genetic material.
Monomeric antibodies are present in the IgG, IgD, IgE isotype classes. IgA and IgM are oligomeric complexes that consist of more individual antibodies.
Monospecific binding implies the specific binding of one epitope, cell or tissue type. The term describes a functional property of antibodies, but does not relate to the origin or production process. Monospecific antibodies may be produced from the same cell clone (i. e. monclonal antibodies) but this is not a necessary condition for monospecific binding.
Neutralization is the effect of so called neutralizing antibodies that bind to pathogens or toxins in a way that renders them unable to cause disease anymore. The antibodies might disrupt a crucial biological process that is necessary for e. g. entry into host cells or a protein-protein interaction with pathogenic downstream effects.
Opsonization is the coating of a pathogen’s surface with antibodies. It is a strong signal for opsonin receptors on phagocytes to encapsulate and digest the opsonized particle. This process is crucial in the defense against infection.
An organism that is pathogenic to humans, i. e. disease-inducing. Examples are found among bacteria, fungi, viruses, parasites, worms, etc. However, not all bacteria, fungi and viruses are pathogenic to humans: some types may cause disease in animals but not in humans.
Phage display makes use of bacteria infecting phages that can be genetically engineered to display antibodies on their surface. They can be selected by their affinity towards their antigen, while uninteresting phages are washed away. The candidates are allowed to infect bacteria to multiply and then be harvested. This process is repeated with more stringent conditions to select high affinity antibodies for virtually any antigen of interest.
Phagocytes are immune cells that fight pathogens and harmful substances by ingesting them and subsequent enzymatic digestion into their building blocks in a process called phagocytosis.
Phagocytosis is a major protective process and a constituent of the immune system. Phagocytes encapsulate pathogenic particles, ingest them and split them into their constituent amino acids, carbohydrates, etc. Phagocyte surfaces contain numerous receptors involved in regulation of phagocytosis. The opsonin receptor recognizes particles that are coated with antibodies (opsonization) and initiates phagocytosis.
Polyclonals which vary in their affinity and specificity towards multiple epitopes of an antigen because they are derived from many clones of B cells. Immunization of animals usually leads to the formation of numerous different antibodies with a broad distribution of specificities.
Post-translational modifications are molecular features of proteins that are attached to it after translation in the ribosome in higher organisms. Most prominent is the N-glycosylation of amino groups, leading to N-glycans. They often play a crucial regulatory role, e. g. afucosylated antibodies.
Purification denoted the cleaning of a raw product during a manufacturing procedure. Depending on the product, impurities, salts, liquid media, solvents, or water must be separated from the desired component. Numerous techniques were developed to accomplish this task: precipitation, extraction, distillation, chromatography, etc.
Recombinant antibodies are antibodies that were produced using recombinant technology, i.e. modern molecular biology and biochemical methods, to engineer genetic material coding for antibodies with desired properties.
Recombinant antibody expression describes the process of antibody synthesis in ribosomes according to a recombinantly designed blueprint in a biological system (expression system).
Recombinant antibody production implies the manufacture of recombinant antibodies on larger scale with the intent to use them to make another product, such as an antibody therapeutic or a diagnostic tool kit.
Recombinant proteins are polypeptides that were produced using recombinant technology to engineer their properties on the genetic level. Recombinant antibodies are a subset of recombinant proteins.
Ribosome display is functionally related to the phage display method. It is used to create antibodies with desired binding properties by changing the genetic blueprint to halt protein expression in the ribosome in a certain way that prevents disassembly of the complex – the ribosome displays the antibody as well as the associated genetic material. This makes the overall process less laborious, as cell culture is avoided.
Specificity describes the preference of a competitive binding interaction between three types of molecules over one another. In a mixture of molecules A, B and C two types of complexes are possible: AB and AC. If the amount of e. g. AB is much higher than AC, then A is said to possess specificity towards B over C.
STR-silenced antibodies are engineered in a way to abolish any signaling through Fc receptors, hence minimizing immune system activation. This is a breakthrough approach to truly Fc-silenced antibodies.
T cells are lymphocytes and key players in the human immune system. Prominent members of this cell class are natural killer cells (NK cells) that fight pathogens. Other types of T cells serve as regulatory cells via modulating the activity of other immune system components.
Therapeutic antibodies are antibodies that have a therapeutic effect when administered to patients. They are used in many distinct diseases like cancers or autoimmune diseases. Recombinant monoclonal antibodies are the most promising representatives due to their high specificities and low degree of immunogenicity, hence minimizing adverse side effects.
Transient antibody production involves introducing antibody genes into cells to produce antibodies for a limited time, without permanently modifying the cell’s DNA.
In molecular biology a vector is any particle (plasmid, virus, phage, etc.) that carries genetic material in a way that is suitable to transfer it into another cell. They are crucial tools to express engineered proteins in biotechnological processes.
Yeast display is a molecular biology method functionally related to phage display. Antibody genes are modified in such a way that the expressed antibodies are attached to the yeast cell’s surface, hence allowing selection of the desired cells for further processing.