Highly sensitive pH-modulated paper-based platform for point-of-care osteopontin quantification in osteoporosis screening

Abstract

This work presents the development of paper-based colorimetric biosensors for the detection of the osteoporosis biomarker osteopontin (OPN), which can be used for point-of-care analysis. For this purpose, simple filter paper has been used, on which test zones are printed using a conventional office desktop printer. The paper was chemically modified with glutaraldehyde to create binding sites for gold nanoparticle-conjugated antibodies. The surface modification was confirmed by Fourier-transform infrared spectroscopy (FTIR). Colorimetric detection was performed using phenol red as a pH-sensitive sensor probe, and this sensitivity was visualised by the immediate generation of brick red color in the test zone of the biosensor upon immunocomplex formation between antibody-conjugated gold nanoparticles and the osteopontin biomarker. The color intensity was quantified using reflectance spectroscopy and the RGB coordinate provided by the ImageJ Program from the photograph taken with a smartphone camera. The sensor demonstrated good noticeable sensitivity in the range of 1–125 ng mL⁻¹ (R² > 0.99) in buffer and in the range of 1–100 ng mL⁻¹ (R² > 0.957) with a detection limit of 0.52 ng mL⁻¹ in buffer and 0.48 ng mL⁻¹ in commercial human sera solution. The detection is possible within 15 minutes of reaction time. In addition, this biosensor shows good selectivity toward other interfering biomarkers, such as bovine serum albumin, osteocalcin, and CTX-1. The developed assay meets the criteria of being easy to use, affordable, and time-saving and requiring a smaller sample volume, which is beneficial for future screening and detection of osteoporosis.