Integrated ratiometric fluorescence probe-based acoustofluidic platform for visual detection of anthrax biomarker

The sensitive detection of dipicolinic acid (DPA) as an excellent biomarker of Bacillus anthracis, especially through visual point-of-care testing, is significant for accurate and rapid diagnosis of anthrax to timely prevent anthrax disease or biological terrorist attack. Herein, an acoustofluidics-based colorimetric platform with the integrated ratiometric fluorescence probe (INT-probe) was fabricated, which improved the sensitivity of visual detection for DPA and overcame the poor reproducibility of the existing acoustofluidics-assisted colorimetric analysis. For the design of INT-probe, Eu³⁺-EDTA complex as sensing moiety was grafted onto the surface of blue organosilane-functionalized carbon dots (SiCDs)-doped SiO₂ nanoparticles (NPs). Upon exposure to DPA, Eu³⁺ was sensitized by DPA to emit red luminescence, while the SiCDs as reference inside the SiO₂ NPs still kept the blue fluorescence unchanged. Attributed to the acoustic radiation force-driven enrichment of the INT-probe, slight color changes caused by low concentration of DPA could be amplified and distinguished by naked-eyes/smartphone. With the increase of DPA concentration, obvious color variations of INT-probe/DPA aggregates from blue to pink could be observed, and the color information of the fluorescent aggregates was converted to red, green and blue values for quantitative analysis, whose lowest detectable concentration reached 100 nM that is about 2-3 orders of magnitude lower than the infectious dosage of Bacillus anthracis spores (60 μM). Importantly, benefiting from the great color signal enhancement by acoustofluidic sensing platform, the usage of Eu³⁺ reduced to as low as 0.273 μmol per gram of SiO₂ NPs, providing a meaningful way to utilize lanthanide resource efficiently.