Intact fibroblast growth factor 23 and fragments in plasma from Gambian children

SUMMARY: Fibroblast growth factor 23 (FGF23) is grossly elevated in Gambian children with rickets and, at a lower prevalence, in those without bone deformities. We used western blotting to mimic the detection capabilities of the C-terminal FGF23 enzyme-linked immunosorbent assay (ELISA). Only intact FGF23 hormone was present in Gambian plasma samples from children with and without rickets. INTRODUCTION: Elevated circulating FGF23 concentrations have been detected in plasma samples from Gambian children using the C-terminal Immutopics ELISA. The Immutopics ELISA detects both the intact FGF23 hormone and the C-terminal fragment. The aim of this study was to determine whether the elevated FGF23 concentrations as detected by the ELISA were predominantly due to a high proportion of intact FGF23 hormone and/or C-terminal FGF23 fragments. METHODS: Stored, frozen plasma samples from previous studies of Gambian children with known concentrations of FGF23 as determined by C-terminal Immutopics ELISA assay, were selected for western blotting analysis: from children with rickets-like bone deformities (n = 4) and local controls (n = 4), with elevated >900 RU/ml (n = 2) and normal <30 RU/ml (n = 2; from each group). The anti-FGF23 polyclonal antibody that recognizes the C-terminal of FGF23 (as used in the Immutopics kit) was used as the primary antibody and the anti-IgG polyclonal antibody conjugated to horseradish peroxidase (HRP) was used as the secondary antibody. RESULTS: Firstly, C-terminal FGF23 fragments, although detectable in standards from the Immutopics ELISA kit, were not in the Gambian plasma samples. Secondly, there was no difference in the size of FGF23 molecules present in plasma from children with rickets-like bone deformities and children from the local community. CONCLUSIONS: Western blotting has provided evidence that elevated FGF23 concentrations, as determined by the C-terminal Immutopics ELISA, measured in Gambian children with and without rickets-like bone deformities was not caused by an increased proportion of circulating inactive C-terminal fragments.