Biomedical Engineering Reference
In-Depth Information
Time . Most bioassays take days, and in some cases week, to run. This can render routine bio-
assays diffi cult, and impractical to undertake as a quick QC potency test during downstream
processing.
Cost . Most bioassay systems, in particular those involving whole animals, are extremely expen-
sive to undertake.
Because of such diffi culties alternative assays have been investigated, and sometimes are used
in conjunction with, or instead of, bioassays. The most popular alternative assay system is the im-
munoassay.
Immunoassays employ monoclonal or polyclonal antibody preparations (Chapter 13) to detect
and quantify the product (Box 7.1). The specifi city of antibody-antigen interaction ensures good
assay precision. The use of conjugated radiolabels (RIA) or enzymes (EIA) to allow detection of
antigen-antibody binding renders such assays very sensitive. Furthermore, when compared with
Box 7.1
Immunoassays
In addition to their therapeutic use (Chapter 13), antibodies are frequently employed as di-
agnostic reagents because they exhibit extreme specifi city in their recognition of a particular
ligand, i.e. the antigen that stimulated their production. Antibody preparations are often used in
the detection and quantifi cation of a wide variety of specifi c analytes, including specifi c thera-
peutic proteins, and assays that employ antibodies in this way are termed immunoassays. The
substance of interest is fi rst employed as an antigen and injected into animals in order to elicit
the production of antibodies against that particular molecule. Either monoclonal or polyclonal
antibody preparations (Chapter 13) may be used in immunoassay systems.
Antibody molecules have no inherent characteristic that facilitates their direct detection in
immunoassays. A second important step in developing a successful immunoassay, therefore,
involves the incorporation of a suitable 'marker'. The marker serves to facilitate the rapid de-
tection and quantifi cation of antibody-antigen binding. Earlier immunoassay systems used ra-
dioactive labels as a marker (radioimmunoassay; RIA) although immunoassay systems using
enzymes (enzyme immunoassays; EIA) subsequently have come to the fore. Yet additional im-
munoassay systems use alternative markers including fl uorescent or chemiluminescent tags.
EIA systems take advantage of the extreme specifi city and affi nity with which antibodies
bind antigens which stimulated their initial production, coupled to the catalytic effi ciency of
enzymes, which facilitates signal amplifi cation as well as straightforward detection and quanti-
fi cation. In most such systems, the antibody is immobilized on the internal walls of the wells in
a multi-well microtitre plate, which therefore serves as collection of reaction mini-test tubes.
Since their initial introduction over 30 years ago many variations on the basic enzyme im-
munoassay concept have been designed. One of the most popular EIA systems currently in use
Search WWH ::




Custom Search