Characterization and use of the antibodies obtained

Antibodies can be separated into different classes IgM, IgG, IgA. and IgE) and subclasses (IgGl, IgG2a, IgG2b, and so on) on the basis of the structure of their heavy chains. This can be achieved using antisera direaed against epitopes on the heavy chains that arc specific for the particular isotype. The different classes have different properties and this may be important in determining the usefulness of the antibodies for example in diagnostic and therapeutic applications. 

Coat plates with monoclonal or polyclonal antibodies (5 p /ml of PCB) using the 

Add 50 pi of test mAb to each well and incubate for 1-4 h at room temperature or 

Wash three times with PBST and determine the amount of radioactivity bound. Include mAbs of known Isotype as controls. 

Occasionally, a mAb will show reactivity with more than one anti-isotype antibody in this capture assay. In this case, try immunoprecipitation in agarose gels (Ouchterlony procedure, ref. 16). Because a crosslinked lattice must be formed between several different epitopes on the antigen and several antibodies to give an immunopredpitate the Ouchterlony procedure gives few false positives. 

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This chapter will provide a detailed protocol to obtain and purify mouse OECs using a specific method to eliminate fibroblasts, to expand them, and, finally, to characterize these cells by means of immunocytochemistry and the use of specific antibodies (Fig. 1). 

There are literally hundreds of markers that are currently available for the mouse and human than can be used to characterize lymphoid and myeloid cells and subsets in primary and secondary lymphoid organs. Many of the markers expressed in mammals are highly conserved across species and have been designated as genetic clusters of differentiation (CD). CDs can be identified with fluorescently labeled monoclonal antibodies. As presented previously, when combined with other fluorescent probes, important information on intracellular biochemistry and cell function can be obtained. Many of the biochemical markers used by immunotoxicologists are common to other... 

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