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- Fang, Xiaotian T., et al.
(författare)
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Efficient and inexpensive transient expression of multispecific multivalent antibodies in Expi293 cells
- 2017
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Ingår i: Biological Procedures Online. - : Springer Science and Business Media LLC. - 1480-9222. ; 19
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Tidskriftsartikel (refereegranskat)abstract
- Background: Immunotherapy is a very fast expanding field within drug discovery and, hence, rapid and inexpensive expression of antibodies would be extremely valuable. Antibodies are, however, difficult to express. Multifunctional antibodies with additional binding domains further complicate the expression. Only few protocols describe the production of tetravalent bispecific antibodies and all with limited expression levels.Methods: Here, we describe a protocol that can produce functional tetravalent, bispecific antibodies at around 22 mg protein/l to a low cost. The expression system is based on the Expi293 cells, which have been adapted to grow in denser cultures than HEK293 cells and gives higher expression yields. The new protocol transfects the Expi293 cells with PEI (which has a negligible cost).Results: The protocol has been used to generate multiple variants of tetra-and hexavalent bispecific antibodies with yields of around 22 mg protein/l within 10 days. All materials are commercially available and the implementation of the protocol is inexpensive and straightforward. The bispecific antibodies generated in our lab were capable of binding to all antigens with similar affinity as the original antibody. Two of the bispecific antibodies have also been used in transgenic mice as positron emission tomography (PET) ligands to successfully detect amyloid-beta (A beta) aggregates in vivo.Conclusions: This protocol is the first describing transfection of the human Expi293 cells with PEI. It can be used to generate functional multi-specific antibodies in high amounts. The use of biological drugs, and in particular multispecific antibodies, is rapidly increasing, hence improved protocols such as the one presented here are highly valuable.
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| 2. |
- Hermansen, Russell A., et al.
(författare)
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Extracting functional trends from whole genome duplication events using comparative genomics
- 2016
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Ingår i: Biological Procedures Online. - : Springer Science and Business Media LLC. - 1480-9222. ; 18
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Forskningsöversikt (refereegranskat)abstract
- Background: The number of species with completed genomes, including those with evidence for recent whole genome duplication events has exploded. The recently sequenced Atlantic salmon genome has been through two rounds of whole genome duplication since the divergence of teleost fish from the lineage that led to amniotes. This quadrupoling of the number of potential genes has led to complex patterns of retention and loss among gene families. Results: Methods have been developed to characterize the interplay of duplicate gene retention processes across both whole genome duplication events and additional smaller scale duplication events. Further, gene expression divergence data has become available as well for Atlantic salmon and the closely related, pre-whole genome duplication pike and methods to describe expression divergence are also presented. These methods for the characterization of duplicate gene retention and gene expression divergence that have been applied to salmon are described. Conclusions: With the growth in available genomic and functional data, the opportunities to extract functional inference from large scale duplicates using comparative methods have expanded dramatically. Recently developed methods that further this inference for duplicated genes have been described.
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