Unusual results from immunoassays and the role of the clinical endocrinologist

Endocrinology was revolutionized in the 1960s by the introduction of hormone measurements based on immunoassay and for quantitative analysis of proteins and peptides there are still few viable alternatives. Before that time the concentrations of hormones were determined by chemical and biological measures that varied in specificity, were tedious and often needed large samples of biological fluid or tissue to enable detection of the hormone. Immunoassays are based on recognition of molecules by antibodies. Polyclonal antibodies are raised in a number of animal species. Monoclonal antibodies in murine systems are widely used. As a label, radioactivity is often now substituted with a fluorescent, chemiluminescent tag or even an enzyme reaction. Not all assay systems depend upon separation of the antibody-bound elements. Immunoassay methods can show such high specificity and sensitivity that there is a sense of security in the value of laboratory results. There are, however, instances where a clinician finds that one or more results make no sense in terms of the patient presentation or overall clinical /endocrine picture. Clinicians must be alert to the fact that immunoassays do not always produce correct results and they will require some appreciation of where problems arise. A test request seeks to refute or confirm a clinical differential and any discrepancies in patient data should be reported to the laboratory so that appropriate investigations can be carried out. Unusual results are largely due to interferences from sample constituents (endogenous factors) and many such phenomena are recognized. These factors can be detected and eliminated, but they vary from patient to patient and also from time to time in one patient. 1 Results higher than real are recorded due to a lack of antibody specificity, although with changes in the formulation of some assays, particularly from manual techniques to highly automated procedures, negative effects can now also be encountered. Interferences caused by external factors are defined as exogenous factors and may be the result of pre-analytical factors, such as sample collection, but results can also be influenced by assay formulation. Rather than disregard any mismatch, there needs to be an explanation for these discrepancies in results to avoid further confusion about patient diagnosis and management and wasted resources over further tests that may be meaningless. Laboratories use numerous quality assurance schemes in their routine testing to identify systematic errors. For effect, these schemes need to be timely. The samples circulated can be selected to address specific analytical issues so as to test ability to identify erroneous results arising from aberrant patient samples. Clinicians can provide valuable materials for these quality exercises. When alerted to assay problems, the laboratory can take steps to derive accurate test results. Any form of interference may result in incorrect values being produced, upon which clinical decisions are made. Many such problems have been reported in the literature. Table 1 includes a number of examples. In some cases there have been medico-legal actions arising from the clinical judgements. Despres and Grant 25 reviewed the clinical consequences of failure to recognize interferences in thyroid assays. Clinicians need to be aware that analysers may not always provide the best measure of the most appropriate hormone. For instance, vitamin D2 therapy may not be detected with certain 25-hydroxy D assays. 26 Although the immunoassay has remarkable flexibility and sensitivity as an analytical tool, its reliability compared with other automated technologies used in biochemical analyses needs attention. 27