Abstract
BackgroundThe ability to demonstrate the reproducibility of gene expression microarray results is a critical consideration for the use of microarray technology in clinical applications. While studies have asserted that microarray data can be "highly reproducible" under given conditions, there is little ability to quantitatively compare amongst the various metrics and terminology used to characterize and express measurement performance. Use of standardized conceptual tools can greatly facilitate communication among the user, developer, and regulator stakeholders of the microarray community. While shaped by less highly multiplexed systems, measurement science (metrology) is devoted to establishing a coherent and internationally recognized vocabulary and quantitative practice for the characterization of measurement processes.ResultsThe two independent aspects of the metrological concept of "accuracy" are "trueness" (closeness of a measurement to an accepted reference value) and "precision" (the closeness of measurement results to each other). A carefully designed collaborative study enables estimation of a variety of gene expression measurement precision metrics: repeatability, several flavors of intermediate precision, and reproducibility. The three 2004 Expression Analysis Pilot Proficiency Test collaborative studies, each with 13 to 16 participants, provide triplicate microarray measurements on each of two reference RNA pools. Using and modestly extending the consensus ISO 5725 documentary standard, we evaluate the metrological precision figures of merit for individual microarray signal measurement, building from calculations appropriate to single measurement processes, such as technical replicate expression values for individual probes on a microarray, to the estimation and display of precision functions representing all of the probes in a given platform.ConclusionWith only modest extensions, the established metrological framework can be fruitfully used to characterize the measurement performance of microarray and other highly multiplexed systems. Precision functions, summarizing routine precision metrics estimated from appropriately repeated measurements of one or more reference materials as functions of signal level, are demonstrated and merit further development for characterizing measurement platforms, monitoring changes in measurement system performance, and comparing performance among laboratories or analysts.
Highlights
The ability to demonstrate the reproducibility of gene expression microarray results is a critical consideration for the use of microarray technology in clinical applications
The documentary standard International Organization for Standardization (ISO) 5725-1 [6] details the basic concepts and estimation techniques for assessing metrological "accuracy" which is defined as a combination of two concepts, "trueness" and "precision." These concepts are formally defined in the Vocabulary of International Metrology (VIM) [7] base document and more cogently described in ISO 3534 [8]: trueness is the closeness of a measurement to an accepted reference value and precision is the closeness of measurement results to each other
The precision of a defined measurement process can be characterized using three nested metrics: "repeatability," "intermediate precision," and "reproducibility." These measures of precision are defined in terms of the conditions that apply when the measurements are obtained, including: operator, equipment, calibrations, environmental conditions, and the period of time between measurements
Summary
The ability to demonstrate the reproducibility of gene expression microarray results is a critical consideration for the use of microarray technology in clinical applications. While studies have asserted that microarray data can be "highly reproducible" under given conditions, there is little ability to quantitatively compare amongst the various metrics and terminology used to characterize and express measurement performance. While shaped by less highly multiplexed systems, measurement science (metrology) is devoted to establishing a coherent and internationally recognized vocabulary and quantitative practice for the characterization of measurement processes. The microarray community has yet to adopt a standardized terminology and practice for characterizing performance that can facilitate clear communication among the user, developer, and regulator stakeholders. The measurement science (metrology) community is devoted to establishing a philosophically coherent terminology and practice for characterizing and communicating measurement performance [4]. While microarrays can generate vastly more data per sample than is typical of the processes that shaped the development of these documents, we believe that this pre-existing metrological framework can be extended to microarrays and other highly multiplexed measurement processes
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