The Nature of Calibrators in Immunoassays: Are they Commutable with Test Samples? Must they Be?

Abstract

Immunoassay systems require calibration protocols that are normally more sophisticated than many analytical techniques in routine clinical use. Calibrators used in such assays may differ significantly from the analyte in clinical specimens. Differences in the properties of calibrators, or reference materials, from those of clinical specimens may include: species origin of the calibrator for an analyte; integrity of the molecular species; matrix of the calibration solution; addition of preservative agents. Owing to the large number of potential differences in the properties of calibrators and those of serum specimens that may affect immunoassay results, the concept of commutability that we originated and first applied to enzyme activity measurements can readily be applied to immunoassay determinations. We specifically examined the properties of calibration materials in nine commercial immunoassay tests for human thyrotropin. Significant non-commutability of materials was demonstrated. The measured results obtained with authentic patient sera differed by a factor of approximately two fold between the techniques exhibiting the lowest and greatest numeric results. Considerably larger intermethod biases were found for calibration materials. Multivariate analysis revealed that the patient sera formed a highly focussed pattern. The calibration materials for one instrument system also focussed in this group. Other calibrators formed three foci indicating similar patterns of commutability within each of the three groups. Clustering was independent of the amount of thyrotropin in the patient specimens, but appeared to be concentration-dependent for at least some of the calibrators. Thus the availability of a common calibration material appears feasible, but not presently available in many commercial products. A processed human serum, a candidate material for use in our proficiency testing program, was projected in the same cluster as authentic patient sera indicating that this material has intermethod properties identical to patient sera (i.e. fully commutable).