Human Monoclonal Autoantibodies
Thomas F. Kresina in Monoclonal Antibodies, Cytokines, and Arthritis, 2020
The hybridoma formation is accomplished through physical fusion of the activated B cell with the fusing partner cell line. Most often, this has been achieved by mixing the cells in the presence of polyethylene glycol (PEG) (18). However, a technology has recently been developed for achieving fusion of cells through electrical breakdown of their membranes (20). In our hands, this “electrofusion” has shown distinct advantages over PEG in that it appears to result in a greater efficiency of fusion and can be used to fuse extremely small numbers of activated B cells (21). Therefore, we currently use electrofusion exclusively for all hybridoma formation. The approach of EBV activation with subsequent fusion by PEG or electrofusion has been effective in the production of human monoclonal antibodies to a variety of antigens (Table 1).
The Inducible System: History of Development of Immunology as a Component of Host-Parasite Interactions
Julius P. Kreier in Infection, Resistance, and Immunity, 2022
Antibodies are protein molecules secreted by B lymphocytes after they have encountered antigen. Each lymphocyte and its clonal descendants are capable of secreting only one type of specific antibody. The invading antigen “selects” the matching clone which then proliferates and differentiates into antibody-secreting plasma cells. Elucidation of the structure of the antibody molecule by analytical chemistry and use of the techniques of molecular biology to study the genome led to the discovery that genes undergo extensive recombination of their DNA in order to generate the remarkable antibody diversity that B lymphocytes possess. Since antigens are complex, many different clones are activated and many different antibodies are produced in an immune response to any antigenic exposure. The artificial production of hybridomas, however, allows for the production of monoclonal antibodies.
Immune system modulators
Gabriel Virella in Medical Immunology, 2019
Monoclonal antibodies represent one of most groundbreaking developments in immunology of the past century. By taking advantage of the ability of B lymphocytes to produce antibodies directed against specific antigens, newer agents are being developed that can have a more selective impact on the immune response. To produce monoclonal antibodies, an animal host (typically murine) is immunized with a specific antigen against which antibodies are to be formed. The mature B cells that develop are then harvested and combined with a line of immortal myeloma cells, forming hybridomas that continually replicate. These hybridomas are then screened to select the lines producing the desired antibody. In order to prevent an immune response or even a hypersensitivity reaction from occurring due to the use of a fully murine monoclonal antibody, efforts have been made to produce genetically modified antibodies that are either chimeric (antibodies that combine the antigen-binding region derived from the murine cells with a human Ig constant region) or fully humanized. These efforts help to decrease the development of antibodies directed against the monoclonal antibody. A number of agents have been developed to assist in the treatment of organ rejection, malignancy, and a variety of different autoimmune conditions.
A natively paired antibody library yields drug leads with higher sensitivity and specificity than a randomly paired antibody library
Published in mAbs, 2018
Adam S. Adler, Daniel Bedinger, Matthew S. Adams, Michael A. Asensio, Robert C. Edgar, Renee Leong, Jackson Leong, Rena A. Mizrahi, Matthew J. Spindler, Srinivasa Rao Bandi, Haichun Huang, Pallavi Tawde, Peter Brams, David S. Johnson
Mouse immunization followed by hybridoma screening has long been used successfully for discovery of therapeutic monoclonal antibodies (mAbs) that are approved by the US Food and Drug Administration.1 Using this method, mice are first immunized with an immunogen and adjuvant. Conventionally, wild type mice are used, but the use of mice transgenically engineered to express human immunoglobulin (Ig) V(D)J sequences has recently become more popular.2 After assessing titer and sacrificing the animals, hybridomas are generated by fusing primary B cells with myeloma cells.3 Although hybridoma protocols are cheap, the process of screening polyclonal pools of hybridomas remains expensive and inefficient. To increase hybridoma screening throughput, large-scale antibody discovery groups, including one at Bristol-Myers Squibb (BMS), use costly robotic systems to automate workflows.
The Antibody Society’s antibody validation webinar series
Published in mAbs, 2020
Jan L.A. Voskuil, Anita Bandrowski, C. Glenn Begley, Andrew R.M. Bradbury, Andrew D. Chalmers, Aldrin V. Gomes, Travis Hardcastle, Fridtjof Lund-Johansen, Andreas Plückthun, Giovanna Roncador, Alejandra Solache, Michael J. Taussig, James S. Trimmer, Cecilia Williams, Simon L. Goodman
On the other hand, many researchers consider a monoclonal antibody to be homogenous by virtue of its production by a monoclonal hybridoma, and assume it to be a single antibody with specificity to one epitope. However, one study showed that about 30% of the hybridomas tested express additional light chains, often derived from the cell-fusion partner, thus rendering the expressed antibodies non-specific.23 Monoclonal antibodies can also be derived from a hybridoma expressing more than one heavy chain allele, or from a hybridoma arising from a fusion to two different B cells, thus producing a mixture of specificities. In either case mono-specificity is lost. But it must be emphasized that the observed cross-reactivity of many monoclonal antibodies is an intrinsic molecular property of the homogeneous IgG molecule, which cannot be purified away. In other words, even a “strictly” monoclonal antibody can still be non-specific.
Animal- versus in vitro-derived antibodies: avoiding the extremes
Published in mAbs, 2021
Andrew R.M. Bradbury, Stefan Dübel, Achim Knappik, Andreas Plückthun
The EURL ECVAM Recommendation on Non-Animal-Derived Antibodies9 reiterated previously stated positions:10–12 the widespread adoption of high-quality recombinant antibodies is essential for improving biological research quality and reproducibility. Unlike almost all other reagents, animal-derived antibodies are for the most part not molecularly defined, and sold for what they purportedly do, rather than what they are, a problem recombinant antibodies can overcome. Once sequence-defined, antibodies become immortal: gene synthesis allows the production of essentially identical reagents ad infinitum, as well as variants, where antibody genes are fused to functional moieties or altered, for additional applications. As antibody genes from hybridomas can now be easily sequenced, the same advantages potentially extend to preexisting monoclonals. Thus, independently of the 3Rs, there are strong scientific arguments for sequence-defined recombinant antibodies, which of course must be of high quality and well characterized.3,11
Related Knowledge Centers
- Antibody
- B Cell
- Electroporation
- Immortalised Cell Line
- Monoclonal Antibody
- Polyclonal Antibodies
- Tissue Culture
- Antigen
- Cell Fusion
- Splenocyte