Osteologie 2019; 28(01): 50-51
DOI: 10.1055/s-0039-1679978
Freie Vorträge Grundlagenforschung
Georg Thieme Verlag KG Stuttgart · New York

Development and validation of a novel ELISA for the detection of intact FGF23 in serum and plasma

A Bitzer
1   Biomedica Medizinprodukte GmbH, Wien
,
J Wallwitz
2   The Antibody Lab GmbH, Wien
,
G Berg
1   Biomedica Medizinprodukte GmbH, Wien
,
G Himmler
2   The Antibody Lab GmbH, Wien
› Author Affiliations
Further Information

Publication History

Publication Date:
05 March 2019 (online)

 

Introduction:

Fibroblast growth factor 23 (FGF23) is a bone-derived hormone, suppressing renal phosphate reabsorption and vitamin D synthesis, and stimulating calcium reabsorption in distal tubules of the kidney. The bioactive intact FGF23 contains 251 amino acids and is glycosylated and phosphorylated. Its activity is mediated by binding to FGFR/Klotho receptor complex at the target cell surface. FGF23 is cleaved between Arg179 and Ser180 to an inactive N-terminal and a C-terminal fragment. Increased serum concentrations of intact FGF23 are a hallmark of renal phosphate-wasting diseases such as ADHR, X-linked hypophosphatemia (XLH), tumor-induced osteomalacia, or autosomal recessive hypophosphatemic rikets.

Methods:

Here, we show the development, characterization and validation of a new intact FGF23 ELISA. Epitopes of both monoclonal antibodies were analyzed by overlapping linear peptides spotted to a microarray and binding kinetics were determined with biolayer interferometry. The assay was validated according to standard quality guidelines regarding its specificity, precision, robustness, accuracy and linearity. Assay performance as well as sample measurements of apparently healthy and diseased human subjects were compared with other commercially available assays.

Results:

The structural epitope of the coating antibody is located in the N-terminal part of FGF23, whereas the horseradish peroxidase labelled detection antibody detects a linear epitope at the C-terminal fragment. Both antibodies bind with high affinity. The sandwich immunoassay generates highly specific signals for human intact FGF23. Accuracy, parallelism, as well as intra- and inter-assay precision are within the standard of acceptance. The correlation of apparently healthy and diseased samples with other assays on the market are quite good, but the incubation times and incubation steps are less and it has a broader overall calibration range.

Discussion:

This well-characterized and fully validated intact FGF23 ELISA can be used for the measurement of human serum and plasma samples and may support further FGF23 research.