Characterization of a sandwich ELISA for the quantification of all human periostin isoforms

• Published in: Journal of clinical laboratory analysis Vol. 32; no. 2
• Main Authors: Gadermaier, Elisabeth, Tesarz, Manfred, Suciu, Andreea Ana‐Maria, Wallwitz, Jacqueline, Berg, Gabriela, Himmler, Gottfried
• Format: Journal Article
• Language: English
• Published: United States John Wiley & Sons, Inc 01-02-2018; John Wiley and Sons Inc
• Subjects: Adult; Aged; Aged, 80 and over; allergy; Antibodies; Antibodies - metabolism; Biomarkers; cardiovascular disease; Cell Adhesion Molecules - blood; Cell Adhesion Molecules - isolation & purification; Cell Adhesion Molecules - metabolism; ELISA; Enzyme-linked immunosorbent assay; Enzyme-Linked Immunosorbent Assay - methods; Epitope Mapping; Epitopes; Female; Humans; Isoforms; Male; Middle Aged; oncology; OSF‐2; osteology; periostin; Peripheral blood; Protein Isoforms - blood; Protein Isoforms - isolation & purification; Protein Isoforms - metabolism; Proteins; respiratory disease; sandwich ELISA; cardiovascular disease; osteology; periostin; sandwich ELISA; respiratory disease; OSF-2; allergy; oncology; ELISA
• ISSN: 0887-8013; 1098-2825
• DOI: 10.1002/jcla.22252

Background
Periostin (osteoblast‐specific factor OSF‐2) is a secreted protein occurring in seven known isoforms, and it is involved in a variety of biological processes in osteology, tissue repair, oncology, cardiovascular and respiratory systems or allergic manifestations. To analyze functional aspects of periostin, or the ability of periostin as potential biomarker in physiological and pathological conditions, there is the need for a precise, well‐characterized assay that detects periostin in peripheral blood.
Methods
In this study the development of a sandwich ELISA using monoclonal and affinity‐purified polyclonal anti‐human periostin antibodies was described. Antibodies were characterized by mapping of linear epitopes with microarray technology, and by analyzing cross‐reactive binding to human periostin isoforms with western blot. The assay was validated according to ICH/EMEA guidelines.
Results
The monoclonal coating antibody binds to a linear epitope conserved between the isoforms. The polyclonal detection antibody recognizes multiple conserved linear epitopes. Therefore, the periostin ELISA detects all known human periostin isoforms. The assay is optimized for human serum and plasma and covers a calibration range between 125 and 4000 pmol/L for isoform 1. Assay characteristics, such as precision (intra‐assay: ≤3%, inter‐assay: ≤6%), spike‐recovery (83%‐106%), dilution linearity (95%‐126%), as well as sample stability meet the standards of acceptance. Periostin levels of apparently healthy individuals are 864±269 pmol/L (serum) and 817±170 pmol/L (plasma) respectively.
Conclusion
This ELISA is a reliable and accurate tool for determination of all currently known periostin isoforms in human healthy and diseased samples.