Microfluidic paper-based analytical aptasensor for fluorometric β-lactoglobulin determination

Herein, we present a new luminescent-based analytical method for sensitive fluorometric determination of a milk allergen, β-lactoglobulin (β-LG), in food products based on a portable, low-cost, and easy-to-use microfluidic paper-based analytical device (µPAD). For this purpose, the developed µPAD consisted on a β-LG specific aptamer immobilized on a cellulose disk, and fluorescent Carbon Quantum Dots (CQDs) bioconjugated with its complementary oligonucleotide (DNA), which worked as a label for the aptamer hybridization and its complementary DNA (CQDs-DNA). In a sample with β-LG, the aptamer preferentially binds to β-LG protein and this leads to release of the CQDs-DNA. After washing the µPAD, where the aptamer is immobilized, detection is achieved by fluorescence intensity decrease. An estimation of β-LG concentration can be calculated thanks to the fluorescence changes, for that, a photo of the µPAD is taken using either a digital camera or a smartphone. The µPAD showed an excellent detection response for β-LG in the range of 1.8 to 500 ng/mL with a limit of detection of up to 0.6 ng/mL using the digital camera. Furthermore, the developed method was successfully applied to the determination of β-LG in food samples such as skimmed milk, crispy rice, cookies and chocolate.