Assay validation and biological variation of serum receptor for advanced glycation end-products

Abstract

Background: The analytical performance characteristics of an enzyme-linked immunosorbent assay for the receptor for advanced glycation end-products (RAGE) were evaluated. The within- and between-subject components of biological variation were also estimated. Methods: Blood was sampled from healthy volunteers into K2-ethylenediamine tetraacetic acid (EDTA) and serum separator tubes (SST) and the stability of RAGE in whole blood, plasma and serum examined. Performance characteristics of the assay were assessed using quality control materials. Three samples were obtained from each of 21 healthy volunteers one-week apart, RAGE measured and components of biological variation estimated. Results: RAGE concentrations in blood specimens collected into K 2-EDTA and SST were stable for at least 6 hours and, after centrifugation, both plasma and serum were stable for at least 24 hours. The RAGE assay had the following characteristics: inter-assay imprecision: coefficient of variation ≤7.3%, working range: 26-5000 ng/L, linearity: r = 0.9977 and detection limit: 26 ng/L. Overall within- and between-subject biological variations were 14.6% and 56.5%, the index of individuality was 0.31 and the reference change value was 49.0% at P < 0.05. Conclusion: Samples for RAGE analyses in serum or plasma can be collected without significant difficulties with the assay showing acceptable analytical performance characteristics. Conventional population-based reference values are of limited utility in diagnosis, indicating that RAGE is likely to be more useful in monitoring disease.