The optimal ionic concentration of sensing buffer in the detection of RNA by using the DNA probe with the silicon...

This study investigates optimal conditions for surface modification and ionic concentration of sensing buffers for miRNA-21 detection using DNA probes on silicon nanowire field-effect transistor (SiNW-FET) biosensors. Ionic strength is a key factor influencing DNA/RNA hybridization efficiency and FET detection sensitivity by affecting duplex formation and the Debye length. An optimal balance between these effects is crucial for ultra-sensitive miRNA detection. The surface functionalization process was optimized through systematic testing of reaction time, temperature, and pH. A 30-min silanization reaction at room temperature without pH adjustment, followed by acetic acid rinsing, resulted in the most uniform silica surface. For hybridization detection, fluorescence microscopy showed that the highest ionic strength (150 mM) of Bis-Tris propane (BTP) buffer produced the greatest hybridization amount. Grazing-incidence small-angle X-ray scattering (GISAXS) confirmed stable secondary structures in DNA/DNA and DNA/RNA hybrids across all ionic strengths. For SiNW-FET measurements, BTP buffers with varying ionic strengths (10 mM, 50 mM, and 150 mM) were tested. A 50 mM BTP buffer provided the optimal balance between ionic strength for hybridization and electric double-layer structure, yielding the highest voltage shifts and enhanced sensitivity for ultra-low miRNA concentrations. Moreover, 50 mM BTP outperformed 50 mM PBS due to BTP's larger counterions reduced ion accumulation on the sensor surface, further improving sensitivity. These findings are crucial for advancing non-invasive liquid biopsy techniques in detecting low-concentration miRNAs.