A two-pronged G-quadruplex construction strategy enables the development of a two-in-one aptasensor for...

Food allergies are a global concern since they threaten human health and are challenging to treat. Avoiding allergen exposure is an effective measure for vulnerable individuals, requiring reliable and simple allergen detection techniques. This can be achieved by integrating aptamer and G-quadruplex (G4) components into a sequence as a smart nucleic acid sensing nanodevice. The challenge lies in the functional structure formation of these components due to the lengthy integrated sequence and its conformational complexities. This study proposes a two-pronged strategy, encompassing both the nucleic acid sequence level and spatial structure levels, to construct a functional parallel G4 for an aptamer-loop-G4 integrated sequence, thereby enabling colorimetric detection of β-lactoglobulin (β-LG). Specifically, through comprehensive tailoring, interfering structures are excluded, and the integrated sequence is modified to adopt an appropriate G4 conformation to enhance signaling efficiency. Additionally, the topologically guided combined use of Mg²⁺ and K⁺ ions coordinates the conformational folding of the aptamer and G4 components, ensuring the recognition capacity of the aptamer. Finally, a β-LG aptasensor is established that presents advantages, such as cost-efficiency, simple operation, 35-min rapid detection, excellent specificity, and sensitivity for a linear range of nearly four orders of magnitude, meeting point-of-care testing (POCT) requirements.