The cell membrane serves as a dynamic interface connecting cells with their environment, harboring biomarkers that mirror mobile identification, standing, and malfunctions [1], [2]. The discharge of mobile contents, reminiscent of nucleic acids, is induced by cell injury. Irregular membrane protein expression is a trademark of circumstances like most cancers and neurodegenerative ailments [3], [4]. Moreover, binding occasions on the interface of immune and tumor cell membranes have important implications for illness development [5], [6]. Consequently, creating applied sciences able to sensitively, precisely, and spatially resolving the biomarkers related to these occasions is paramount for advancing biomedical analysis and bettering scientific follow.
DNA-encoded logic gate programs have just lately emerged as a robust instrument in biosensing for mobile occasion imaging, as a result of their programmability, biocompatibility, versatile molecular design, and ease of integration with purposeful nucleic acids possessing focusing on recognition (aptamers) and catalytic exercise (DNAzymes) [7], [8], [9], [10], [11], [12], [13], [14]. Mixed with DNA monomer-encoded cascade amplification methods reminiscent of hybridization chain response (HCR) [15], [16], [17], [18], [19], rolling circle amplification (RCA) [20], [21], [22], [23], [24], and DNAzyme-catalyzed substrate cleavage [25], [26], [27], [28], [29], DNA-encoded logic gate programs with sturdy capabilities for processing advanced organic alerts and recognizing multidimensional info in multi-target identification, and have been broadly utilized within the delicate and exact detection of disease-related occasions localized on the cell membrane [30], together with monitoring native acidity [31] and enzyme exercise [32], recognition of abnormally expressed cell membrane proteins [33], [34], [35], glycoproteins [36], [37] and glycoRNA detection [38], [39], [40].
Regardless of these advances, there are nonetheless important challenges in attaining exact and delicate multi-target recognition on membrane interfaces for cell occasion evaluation. Concerning multi-target exact recognition, all at the moment reported strategies depend on monovalent logic components for goal recognition and computation [41], [42], [43], [44], [45], [46], [47], [48]. These conventional strategies undergo from low recognition effectivity, poor anti-interference functionality, and a powerful dependence on the spatial distance between heterogeneous targets, which collectively restrict the computational capability of DNA logic gates and hinder their development towards exact multi-target recognition. Due to this fact, setting up a brand new technology of high-performance logic gate components for exact multi-target recognition on the membrane interface is essential to overcoming the bottlenecks within the additional improvement and broad utility of DNA logic gates.
Moreover, DNA-encoded logic gate programs, in exploring high-sensitivity detection of cell membrane native targets that may mirror mobile occasions, usually improve alerts utilizing unconfined, diffusion-limited cascade reactions in answer [33], [34], [35], [36], [37], [38], [39], [40]. Nevertheless, this method inherently faces a number of challenges: it requires excessive concentrations of monomers to keep up response effectivity, reveals gradual kinetics with extended operation occasions, is inclined to non-specific interference in advanced organic environments, and freely flowing in vivo with out being retained on the goal detection web site, limiting its sensible utility potential in vivo. Though the broadly used interface spatially confined sign amplification methods maintain promise for addressing this concern, most research thus far have targeted on stable materials interfaces [49], [50]. DNA amplification programs confined particularly to cell membrane interfaces are uncommon; to our information, no multi-target in-situ imaging research have reported DNAzyme-driven environment friendly sign amplification based mostly on membrane-confined methods.
To beat these limitations, we current an modern platform that permits spatially confined sign amplification on the membrane interface, pushed by built-in DNA logic gates, for imaging a number of mobile occasions. Firstly, we suggest a novel design impressed by the idea of “integration to boost efficiency” from digital units. We innovatively encoded monovalent conventional logic gate components (tLGE)—comprising aptamers for receptor recognition and logic sequences for AND-gate configuration—into multivalent tandem architectures by branched DNA nanowires, thus designing a brand new technology of built-in DNA logic gate components (iLGE). This considerably enhances the DNA logic gates’ multi-target recognition and computational capabilities on the membrane interface. We subsequent constructed a DNA walker machine (DWD) anchored to the cell membrane by way of a cholesterol-modified DNA framework that integrates the DNAzyme sign swap with its fluorescent reporter substrate (Sub). By exactly colocalizing each parts throughout the framework and on the membrane, our twin confinement technique markedly boosts sign amplification effectivity and spatial precision. The AND-gate configuration fashioned by the iLGE and connector prompts the DNAzyme-based amplification cascade by competitively displacing the Block strand within the DWD. Thus, we established a complete platform for the extremely delicate and exact imaging of numerous membrane-localized stimuli (e.g., nucleic acids, proteins, and cells), facilitating the evaluation of mobile occasions starting from injury and oncogenesis to NK-tumor cell interactions (Fig. 1). This platform advances the examine of cell membrane native biomarkers and holds promise for early illness prognosis and remedy analysis.
