25(OH)-Vitamin D Assay in Plasma: Experience in Using a Commerical Kit Assay for Survey Work

Abstract

Measurement of plasma 25(OH)-vitamin D (250HD) is the accepted index of vitamin D status for people with normal vitamin D metabolism. 1 Although several assay methods for plasma 250HD, based on high-performance liquid chromatography, or Sephadex separations combined with competitive protein binding have been reported, there remains a need for a well-characterized, widely accepted procedure which has a high potential throughput, and long-term reliability. This report describes the precision and long-term reliability of a commercial kit assay during a survey of 737 British children aged 1·5 to 4·5 years, over a period of 1 year. 2 Population survey work in particular, requires precise measurements in both the normal and subnormal ranges; freedom from long-term drift, and freedom from stepwise changes in sensitivity. The commercial kit assay used was that based on the procedure of Hollis et al., 3and marketed by Incstar Corporation (Stillwater, Minnesota, USA). It employs a specific antibody, raised in goats against a 250HD derivative, and a radiolabelled 250HD as tracer, which was originally 3H, but now uses 1251. For convenience, each kit was subdivided into two runs, for I day's work. In each assay 12 unknowns were analysed in duplicate and a set of control sera were placed at the beginning and at the end. The assay was performed according to manufacturer's instructions except that two separate aliquots of standard plasma (or QC) sample were extracted and assayed once, instead of performing a single extraction to generate two measurements, as suggested in the instructions. Radioactivity estimation and data-reduction were performed with a Wallac (Turku, Finland) Wizard -y-counter,