miRNA purification with an optimized PDMS microdevice: Toward the direct purification of low abundant circulating biomarkers

• Published in: Biophysical chemistry Vol. 229; pp. 142 - 150
• Main Authors: Santini, G.C., Potrich, C., Lunelli, L., Vanzetti, L., Marasso, S.L., Cocuzza, M., Pirri, F.C., Pederzolli, C.
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.10.2017
• Subjects: Biomarkers - blood; Circulating microRNAs; Dimethylpolysiloxanes - chemistry; Fluorescent Dyes - chemistry; Humans; Isocyanates - chemistry; Microfluidic Analytical Techniques - instrumentation; Microfluidic Analytical Techniques - methods; MicroRNAs - blood; MicroRNAs - chemistry; MicroRNAs - isolation & purification; Microscopy, Atomic Force; On-chip analysis; On-chip purification; PDMS microdevice; Photoelectron Spectroscopy; Real-Time Polymerase Chain Reaction; Silanes - chemistry; Surface Properties; On-chip analysis; Circulating microRNAs; PDMS microdevice; On-chip purification
• ISSN: 0301-4622; 1873-4200
• DOI: 10.1016/j.bpc.2017.04.009

A reliable clinical assay based on circulating microRNAs (miRNAs) as biomarkers is highly required. Microdevices offer an attractive solution as a fast and inexpensive way of concentrating these biomarkers from a low sample volume. A previously developed polydimethylsiloxane (PDMS) microdevice able to purify and detect circulating miRNAs was here optimized. The optimization of the morphological and chemical surface properties by nanopatterning and functionalization is presented. Surfaces were firstly characterized by atomic force microscopy (AFM), X-ray photoelectron spectroscopy (XPS), fluorescamine assay and s-SDTB (sulphosuccinimidyl-4-o-(4,4-dimethoxytrityl) butyrate) assay and subsequently tested for their capacity to adsorb a fluorescent miRNA. From our analysis, modification of surface charge with 0.1% APTMS ((3-Aminopropyl)trimethoxysilane) and 0.9% PEG-s (2-[Methoxy-(polyethyleneoxy)propyl]trimethoxysilane) performed at 60°C for 10min was identified as the best purification condition. Our optimized microdevice integrated with real-time PCR detection, was demonstrated to selectively purify both synthetic and natural circulating miRNAs with a sensitivity of 0.01pM.