Point: Legitimate and Illegitimate Tests of Free-Analyte Assay Function
| Autor: | John E.M. Midgley, N D Christofides |
|---|---|
| Rok vydání: | 2009 |
| Předmět: |
Analyte
Chromatography Illegitimacy Globulin biology medicine.diagnostic_test Chemistry Biochemistry (medical) Clinical Biochemistry Albumin Ultrafiltration Reproducibility of Results Clinical Chemistry Tests Blood proteins Immunoassay Medical Laboratory Science biology.protein medicine Humans Equilibrium dialysis |
| Zdroj: | Clinical Chemistry. 55:439-441 |
| ISSN: | 1530-8561 0009-9147 |
| DOI: | 10.1373/clinchem.2008.116525 |
| Popis: | In the history of clinical immunoassay, even up to the present day, no group of tests has been subjected to more scrutiny and controversy than analog-type free-analyte assays (1)(2)(3)(4). Unfortunately, inappropriate design of many experiments has led users to an incorrect perspective of how analog assays work. An analysis of the important series of studies by Nelson and coworkers (5)(6)(7)(8)(9)(10) shows that most experiments were conducted outside the valid working limits of analog-type or 2-step free-analyte assays. Here we explore the legitimacy of this viewpoint. In gold-standard methods of equilibrium dialysis and ultrafiltration, free analyte is first separated from that bound to serum proteins. Only then is the free analyte measured by what essentially is a high-sensitivity total analyte assay. For these methods, testing of assay parameters such as recovery of added analyte, linearity of response, and cross-reactivity are legitimate, applying equally to total analyte measurements. When the antibody probe is in direct contact with serum, however, the situation is radically different. For bound thyroxine (T4), serum contains a low-concentration, high-affinity binding protein (thyroxine-binding globulin) and high-concentration, low-affinity proteins (transthyretin and albumin). Free and total analyte are nonlinearly related (except with zero-thyroxine-binding globulin serum) (11). Recovery and linearity experiments in such conditions are thus not feasible, becoming increasingly less accurate at higher thyroxine-binding globulin concentrations (11). Analog-type and equilibrium dialysis/ultrafiltration methods must be calibrated differently. With gold-standard assays, direct calibration is done gravimetrically with known quantities of analyte, a method that is not possible with analog assays. Analog-type methods are calibrated by secondarily applying the values originally obtained from gold-standard methods to the … |
| Databáze: | OpenAIRE |
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