0.001 percent, equivalent to 50 to 500 parasites per microliter of blood), with RIA techniques being slightly more sensitive on average. In general, therefore, the limits of detection of immunoassay techniques approach that of microscopy (which is able to discern 10 to 20 parasites per microliter of blood). The value of RIA methods for both immunoassays and probes (see Genetic-Probe Assays, below) has in the past been to verify that laboratory tests can be developed by using certain combinations of reagents and malarial antigens. However, because RIAs employ radioisotopes that have short shelf lives and are costly and potentially dangerous, they are impractical for routine clinical use, even at the central-hospital level in the developing world. In addition, laboratory personnel have considerably more experience and training in the application of ELISA methods, especially since the start of the AIDS epidemic. Although diagnostic ELISA kits have been used successfully at many well-equipped central health care facilities and in support of epidemiologic or vaccine programs, they are not suited for use in outlying areas where equipment is lacking. The minimum of two to four hours needed to perform an ELISA, and the desirability of testing large numbers of specimens at one time, means that same-day results may not always be available. This is not, however, a problem in large epidemiologic studies, in which same-day results are rarely required.
Antigen Inhibition and Competition In antigen-inhibition and antigen-competition assays, parasite antigens are coated on a surface, such as a microwell plate. The test specimen is centrifuged, and the red blood cells are washed and lysed to extract parasite antigen. The specimen is then mixed with a high-titer serum or monoclonal antibody and placed in the well containing test antigen. If the specimen contains parasite antigens, they will bind to the antimalaria antibodies, thus preventing the antibodies from binding to the antigens on the plate. After rinsing, the amount of antibody bound to the plate is determined by standard RIA or ELISA techniques (Khusmith et al., 1987).
Antigen Capture In antigen-capture (antibody-sandwich) assays, polyclonal or monoclonal antibody is adsorbed to a tube or microwell plate. The specimen is placed into the test vessel and, after washing, a second antibody source conjugated with an enzyme or radioactive label is added. If malarial antigens are present, they serve as a bridge between the antibodies in the tube or well of the plate and the labeled antibodies. The plate is then processed by RIA or ELISA as described above, and the amount of labeling is quantified. Antigen-capture assays may prove to be more practical than antigen-competition assays for diagnosing malaria. Unlike competition assays, capture methods do not require purified malarial antigens and have a greater potential for detecting a broader range of antigens. In