Electrokinetic Preconcentration and Label-Free Electrical Detection of SARS-CoV-2 RNA at a Packed Bed of Bioconjugated Microspheres

In this communication, we demonstrate the electrical detection of SARS-CoV-2 RNA at low femtomolar concentrations without labels or amplification reactions. Following its extraction from virus particles, the viral RNA was electrokinetically preconcentrated (100-fold) within a packed bed of probe-modified microbeads. This preconcentration was accomplished by counter-flow focusing of the RNA along an electric field gradient generated by faradaic ion concentration polarization (fICP). Hybridization of the 30 kb target RNA to the probe-modified beads sufficiently altered their surface charge to yield a measurable change in the ionic conductivity of the packed bed─a feature leveraged for electrical detection. When a single-stranded DNA probe was used, the sensitivity of this enrichment and sensing scheme was low picomolar. However, the utilization of an uncharged PNA probe improved the limit of detection to 3.4 × 10⁶ viral copies/mL (22.5 fM SARS-CoV-2 RNA). These results are significant for three reasons. First, the sensitivity is remarkable, given the micrometer scale of both the beads and interstitial spaces. Additional gains in enrichment and sensitivity are anticipated as fundamental parametric studies and optimization are undertaken. Second, this study reveals the impact of the probe type on the sensitivity of microscale surface ion conduction (μSIC) sensors. Third, the RNA sensing approach has practical advantages including its utilization of off-the-shelf beads, a reagent-free approach, nonoptical readout, and low driving voltage, which render it amenable to point-of-care (POC) implementation.