Antibody construction

The use of a 500-fold molar excess of 2-mercaptoethylamine over the concentration of antibody presence was found to result in a partially reduced antibody in which two disulfides were reduced to yield four thiols (Sun et al., 2005). This strategy can be used to retain a biospecific antibody construct for subsequent discrete conjugation at the hinge region between the heavy chains. 

Sastry, L., Alting-Mees, M., Huse, W.D., Short, J.M., Sorge, J.A., Hay, B.N., Janda, K.D., Benkovic, S.J., and Lerner, R.A. (1989) Cloning of the immunological repertoire in Escherichia coli for generation of monoclonal catalytic antibodies Construction of a heavy chain variable region-specific cDNA library. Proc. Natl. Acad. Sci. USA 86, 5728-5732. 

C. Bourdillon, C. Demaille, J. Moiroux, and J.M. Saveant, From homogeneous electroenzymatic kinetics to antigen-antibody construction and characterization of spatially ordered catalytic enzyme assemblies on electrodes. Acc. Chem. Res. 29, 529-535 (1996). 

FIGURE 5.26. Antigen-antibody construction of a monolayer glucose oxidase electrode with an attached ferrocenium cosuhstrate and cyclic voltammetric response in a phosphate buffer (pH 8) at 25°C and a scan rate of 0.04 V/s. a Attached ferrocene alone, h Addition of the substrate, c Primary plots, d Secondary plot. The numbers on the curves in parts h and c are the values of the substrate concentration in mM. Adapted from Figure 2 in reference 24, with permission from the American Chemical Society. 

Step-by-Step Antigen-Antibody Construction of Multimonomolecular Layered Enzyme Coatings... 

FIGURE 5.28. Antigen-antibody construction of a multilayered enzyme electrode (see the text). Adapted from Figure 2 of reference 26b, with permission from the American Chemical... 

Hudson PJ. Recombinant antibody constructs in cancer therapy. Current Opinion in Immunology 11 548-557 (1999). 

Recombinant technology can be used to produce novel antibody constructs. The potential to produce antibody fragments consisting only of variable regions, or in extreme case of single chain constructs on which the hypervariable regions are assembled has been explored. Protein engineering can also be used to produce... 

Mack, M., Riethmuller, G., and Kufer, P. (1995) A small bispecific antibody construct expressed as a functional single-chain molecule with high tumor-cell cytotoxicity. Proc. Natl Acad Sci USA 92, 7021—7025... 

Pietersz, G. A and McKenzie, I. F. C (1995) The genetic engineering of antibody constructs for diagnosis and therapy, in Monoclonal Antibodies The Second Generation (Zola H., ed.), BIOS Scientific, Oxford, UK, pp 93—117... 

Of all of the potential applications of mAh therapy, cancer treatment has been the most intensively studied. A broad array of antigenic targets and antibody constructs have been evaluated since the advent of mAh technology in 1975, including two decades of clinical trials. Despite... 

Figure 10.17 Phage display of antibodies construction of the genetic information for the displayed Vh-Vl antibody library.

PIMENTEL, G.J., et al., Dimeric scEv antibody construct of ior-CEAl as potential agent for therapeutic apphcation . Trends in Radiopharmaceuticals (Proc. Int. Symp. Vienna, 2005), IAEA, Vienna (2007). 

The first step in the production of monoclonal antibodies is generation of the expression construct. For antibody generation, the antibody construct depicted in Fig. 3.4 has mostly been used. Here, expression of both the tight chain and the heavy chain genes is driven by a cytomegalovirus (CMV) promoter that has been modified to... 

Amplification of the number of copies of expression vector can lead to an increase in productivity of the cell line, but this is not the only method for generating high-yielding cell lines. The GS system [1] and some variants of the DHFR system [23] do not rely upon amplification to achieve high productivities. Instead, these systems rely upon insertion of the antibody construct into a transcriptionally active region to achieve high productivities. Specific production rates for non-amplified antibody-... 

Based on their physiological role, unmodified MAbs can trigger dilTerent immunological reactions that are used in medical applications [55]. These reactions may vary from passive immunization through effector functions to selective cytotoxic effects with bispecific antibody constructs that re-crait cells of the immune system to destroy identified targets [56]. 

Attempts have also been made to use bispecific antibodies to recrait effector cells. A number of challenges need to be overcome in this field. First, a robust method must be found by which such proteins can be produced. Initially, the co-expression of two antibodies in one cell and the separation of the one desired out of the 10 conceivable molecular forms did not seem an attractive proposition. However, a multitude of methods have been reported over the last few years [103, 150] to create bispecific formats of the antibody with a defined molecular composition. These include (1) the co-expression of heavy chains that have been engineered to allow only the desired pairing, (2) the direct chemical linkage of two different Fab fragments, (3) a number of different bispecific recombinant antibody constructs based on scFv fragments fused to heterodimerizing pep-... 

Els Conrath K., Lauwereys M., Wyns L., and Muyldermans S., Camel single-domain antibodies as modular building units in bispecific and bivalent antibody constructs, J. Biol. Chem., 276(10), 7346-7350, 2001. 

Antigen-Antibody Construction of an Immobilized Enzyme Monolayer and Kinetic Analysis of the Enzymatic Catalytic Reaction... 

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