Monoclonal antibodies against antigenic sites

It is entirely possible that surface staining cannot be accomplished before fixation. Some antibody/receptor complexes cannot withstand chemical fixation and/or permeabilization. This is an empirical evaluation of surface, cytoplasmic, or nuclear antigen/ receptor sites, and each must be evaluated before staining or fixation can be accomplished. In this example I, first stain with a monoclonal antibody against a cell surface receptor, fix the cells with ethanol then stain the DNA, and analyze the cells for two colors of fluorescence, red (DNA) and green (surface marker). This approach works for antibody-antigens that are unaffected by fixation. 

When an antigen is injected into an animal, the resulting antibodies are polyclonal, being synthesized by a mixture of B cells. Polyclonal antibodies are directed against a number of different sites (epitopes or determinants) on the antigen and thus are not monospecific. However, by means of a method developed by Kohler and Milstein, large amounts of a single monoclonal antibody specific for one epitope can be obtained. 

Ceriani, R. L., Peterson, J. A., Lee, J. Y., Moncada, R. and Blank, E. W. 1983. Characterization of cell surface antigens of human mammary epithelial cells with monoclonal antibodies prepared against human milk fat globule. Somat. Cell Genet. 9, 415-427. Christie, W. W. and Wooding, F. B. P. 1975. The site of triglyceride biosynthesis in milk. Experientia 31, 1445-1447. 

The monoclonal antibodies used as immunosuppressive agents in tissue transplantation include muromonoab-CD3, daclizumab and basiliximab. Muromonoab-CD3 binds to a specific site on CD3 receptors and interferes with the ability of the TCR to bind the antigen and also inhibits CD3 receptor-dependent signal transduction mechanisms, all of which result in immune suppression. Both daclizumab and basiliximab are monoclonal antibodies directed against IL-2 receptors and consequently inhibit IL-2-dependent responses after tissue transplantation, resulting in immune suppression. The monoclonal antibodies used as immunosuppressive agents are described in detail in Chapter 5. 

Bispecific monoclonal antibodies are artificially developed antibodies with antigenbinding sites physically linked to different specificities. It is thought that bispecific monoclonal antibodies activate the cellular immune response by crosslinking immune cells to tumor cells, thus circumventing the proper structures for tumor cell-immune cell interactions (Koelemij et al., 1999). These antibodies are effective in low concentrations in vivo. For example, Kufer et al. (1996) have combined the anti-CD3 specificity directed against T cells in a bispecific monoclonal antibody, with the specificity against the tumor-associated 17-1A antigen. This antibody could be a major improvement, for example, in the therapy for disseminated micrometastatic tumor cells. 

A sandwich assay for HCG using GOD as label is also described (309). The procedure is based on an amperometric enzyme immunoassay with an electrode-immobilized antibody. The antibody electrode (activated glassy carbon) is used both to separate the assay and to monitor the activity of the bound enzyme label. Two monoclonal antibodies directed against different antigenic sites are used ... 

For a monoclonal antibody, the precursor nucleotide sequence with spaces between codons and translation and with numbers per line CDR-IMGT the origin of each chain sites of disulfide-bridges Ig class and subclass name/structure of the antigen against which the monoclonal antibody is directed. 

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