Immunologic techniques monoclonal antibodies

The assessment of DNA adducts may provide a sensitive indicatCH of previous exposure. The enzyme-linked immunosorbent assay (ELISA) has a lower limit of detection of about 0.08 femtomol per microgram of DNA (Perera et oL, 1982). This assay requires (1) the development of an antibody specific for a certain chemical metabolite bound covalently to DNA and (2) the isolation of DNA from some tissue sample, ag., skin biopsy, or lymphocytes of an exposed individual. It is anticipated that further refinement of such immunologic techniques may lower the threshold of sensitivity by one or two orders of magnitude. One such refined test is the ifitrasensitive ymatic radioimmunoassay (USE-RIA), purported to be about five times more sensitive than ELISA (Hsu et oL, 1981 Shamsuddin et oL, 1985 Harris et oL, 1985). Quantification by the development of monoclonal antibodies to aflatoxin Bj metabolites bound to DNA (Groopman et oL, 1982 Sizaret et oL, 1982) has now been reported. 

In monoclonal antibody purification, biological risks are primarily related to the host animal cells, but also to animal supplements for culture medium such as fetal bovine serum or pure proteins (e.g., bovine albumin, insulin, and transferrin). A special risk associated with production of antibodies with rodent cell lines is their high load of C-type particles. These particles are considered as incomplete retroviruses. The danger regarding infecting humans is not clear. Thus, the efficient separation of these particles must be guaranteed. These particles are quantified either by immunological techniques or electron microscopy. 

Isolation in cell culture and direct visualization by electron microscopy, followed by immunological identification using immunohistochemical techniques, may help if the identity of the VHP agent is unknown. In addition, immunohistochemical techniques, using formalin-fixed tissues, can retrospectively identify specific viral antigens using batteries of specific immune sera and monoclonal antibodies (48). 

Very recently, the preparation of monoclonal antibodies as a highly sophisticated approach to identify micro amounts of pellicle components has been described [43, 52], Immunologic techniques are particularly suitable for identifying small amounts of antigenic components that provide sufficient immuno-genicity to stimulate and induce antibody production by the immune system. Multiple encounters of the immune system with certain antigens amplify the... 

Examination of Pmg antigens by the production of a library of monoclonal antibodies revealed that less than a dozen antigens account for essentially the total immunological response to a mixture of Pmg extracellular glycoproteins. This response can, however, be modified by immunological techniques that suppress antibody production to particular antigens. Even with these relatively sophisticated tools, it has proven difficult to screen... 

For triazine herbicides a very selective and sensitive immunological technique is recommended. The triazine herbicides are covalently bound to soil humic acids. A sandwich-immunoassay based on both polyclonal humic acid-antibodies and monoclonal triazine-antibodies111 was used for triazine detection. The schematic view of sandwich immunoassay is presented in Figure 4.3. The technique is very selective and sensitive. It can assure the best reliability of the analytical information because it does not require a prior separation. 

Production of antisera does not require specialized immunological techniques and cell culture facilities. Therefore it may be a simpler, faster and cheaper way than production of monoclonal antibodies (see section 3.3.). However, it must be noted that antisera-based immunoassays are sometimes less selective than monoclonal antibody-based immunoassay because of the polyclonality of antibodies in the antisera, and good antisera are not reproduced. The production of high affinity antisera may be helped by long-term immunization schedules from the mechanism of antigen-dependent B lymphocyte differentiation and selection (23). 

Immunological techniques For production of monoclonal antibodies and immunoassays standard techniques were applied. 

Immunological detection of ciguatoxins and related polyethers has received particular attention compared to other marine toxins. The initial RIA and enzyme immunoassay employing a polyclonal sheep anti-ciguatoxin antibody revealed cross-reactivities between ciguatoxins and other polyether toxins, suggesting the need for monoclonal antibodies. The availability of monoclonal antibodies allowed for the development of stick enzyme immrmoassay methods and solid-phase immunobead techniques (known as the paddle test), which successfully recognized toxins attached to correction fluid-coated bamboo sticks or paddles previously exposed to toxic fish tissues. 

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