Elsevier

Clinical Biochemistry

Volume 37, Issue 10, October 2004, Pages 867-874
Clinical Biochemistry

Analytical and clinical validation of the 25 OH vitamin D assay for the LIAISON® automated analyzer

https://doi.org/10.1016/j.clinbiochem.2004.06.006Get rights and content

Abstract

Objective: Methods to assess serum 25 OH vitamin D have improved in accuracy, precision, and ease of use. We describe the analytical and clinical validation of an automated, antibody- and microparticle-based, chemiluminescent immunoassay method for the determination of 25 OH vitamin D.

Design and methods: The LIAISON® 25 OH Vitamin D assay is a rapid automated method with first results available in 40 min, and a subsequent throughput of 180 samples per hour. Assay performance characteristics of precision and recovery were determined according to the National Committee for Clinical Laboratory Standards (NCCLS) protocols. Analytical and functional sensitivity were determined according to standard protocols. Samples for method comparison studies were obtained from routine clinical samples submitted for 25 OH Vitamin D determination or from apparently healthy normal volunteers.

Results: The detection limit for this assay was <2.0 nmol/L across three lots of materials. Functional sensitivity (inter-assay imprecision <20%) was 17.5 nmol/L. Total imprecision (CV) was <15% at 42.5–137.5 nmol/L. Mean (SD) recovery was 101% (13%). The assay was linear on dilution. Comparison with radioimmunoassay (RIA) yielded acceptable correlation (r = 0.88) and clinical equivalence in the range from 37.5 to 150 nmol/L.

Conclusions: The LIAISON® 25 OH Vitamin D assay is a rapid, accurate, and precise tool for the measurement of 25 OH vitamin D.

Introduction

Low vitamin D status is being associated with a growing number of diseases. Severe vitamin D deficiency has long been associated with rickets in children and osteomalacia in adults [1], [2]. Serum concentrations of 25 OH vitamin D of less than 40–50 nmol/l (16–20 μg/l) can lead to secondary hyperparathyroidism and its deleterious consequences on bone mass over the long term [3], [4]. Serum 25 OH Vitamin D concentrations less than 80 nmol/L have been shown to impair absorption of intestinal calcium [5]. Vitamin D deficiency has been associated with disturbed muscle metabolism [6], particularly impaired intracellular calcium metabolism [7]. High circulating concentrations of NT-proANP, a marker of the severity of congestive heart failure, has been associated with low circulating 25 OH vitamin D concentrations [8]. In addition, vitamin D status has been linked to prostate, breast, and colon cancer [9], [10], [11]. Currently, the circulating 25 OH vitamin D concentration is the best available index of vitamin D nutritional status [12], [13]. Therefore, the measurement of 25 OH vitamin D is becoming increasingly important to the diagnosis and management of numerous diseases.

The measurement of 25 OH vitamin D has evolved through several technologies. Several high-performance liquid chromatography (HPLC) methods have been described that are labor intensive, often technique dependent, require costly equipment and large sample volumes [14], [15], [16], [17]. Many assays were developed utilizing the naturally occurring vitamin D binding protein from various species and requiring extraction or chromatographic purification as pretreatment steps [18], [19], [20], [21]. These assays often suffered from significant interferences and cross-reactivities with endogenous vitamin D metabolites. Binding protein assays were generally replaced by radioimmunoassays (RIAs) utilizing either 3H- or 125I-labeled tracers [22], [23]. Recently, automated chemiluminescence-based assays have become available, utilizing either the vitamin D binding protein [24], or a specific antibody [25] in the measurement of serum 25 OH vitamin D concentrations. These assays provide the potential to make vitamin D assessment more accessible to the general practitioner. Validation of assay methodologies is mandatory for the modern clinical laboratory. The present study is designed to provide a baseline for assay validation studies for the LIAISON® 25 OH Vitamin D assay [25].

Section snippets

Analytical and clinical samples

Analytical and clinical samples were obtained from normal healthy adult volunteers or hospital outpatients after informed consent or as repository samples from reference laboratories. Samples were analyzed at three separate sites. The two clinical sites utilized one instrument each, while the analytical site utilized four instruments.

Vitamin D assays

Vitamin D was determined in samples with either of two commercially available 25 OH Vitamin D assays. A radioimmunoassay (DiaSorin, Stillwater, MN) was performed

Assay validation

The minimum detectable concentration, defined as the concentration at two standard deviations from the mean zero standard signal, was determined for three lots. The values determined for each of the three lots were 4.3, 4.3 and 4.5 nmol/L, respectively (data not shown). Therefore, the minimum detectable concentration of the assay was less than 5.0 nmol/L.

Since the minimum detectable concentration was less than the lowest reportable limit of the LIAISON® assay, a functional sensitivity was

Discussion

Analytical and clinical method validation is an important requirement as the methods of assessment of 25 OH vitamin D move from labor and equipment intensive, manual extraction methods in the specialty laboratory to the automated non-extraction methods in the general clinical laboratory. Often the laboratory is provided with only minimal or selective comparison data. The data presented here are intended to provide a rigorous baseline for the clinical laboratory in the evaluation of validation

Acknowledgements

Funding for this work was provided by DiaSorin Inc., Stillwater, MN.

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