QCM biosensor based on silver aggregates and DNA ligase for ultrasensitive detection of BRAF-V600E single mutant gene in melanoma.

The detection of BRAF-V600E gene mutations plays a pivotal role in the diagnosis, prediction, and therapeutic intervention of melanoma. In this work, a quartz crystal microbalance (QCM) biosensor was developed to detect the BRAF-V600E point mutation. The gold electrode of the QCM was incubated with a thiol (-SH) modified capture probe, subsequently treated with polyethylene glycol (PEG) to obstruct nonspecific binding sites on the electrode. Subsequently, the target sequence BRAF-V600E (alternatively referred to as BRAF) along with the detection probe were incubated to construct a functionalized layer. T DNA ligase was introduced to facilitate the connection between the detection and capture probes, thereby forming either elongated or abbreviated DNA chains. Ultimately, silver aggregates (AgAs) were formed and bound to DNA bases and exert a significant influence on the sensor's frequency. In instances where the target sequence is BRAF-V600E, the frequency shift is markedly higher in comparison to BRAF, thereby facilitating a clear distinction between the two. The developed biosensor exhibited a linear detection range of 1 pM-10 nM for BRAF-V600E, with a detection limit of 0.73 pM and an R value of 0.9923. The accuracy of the QCM method was rigorously validated through a comparative analysis with quantitative polymerase chain reaction (qPCR). This study holds significant implications for the early detection and precise of melanoma.