Fluorescent molecular probes have become valuable tools in protein research; however, the current methods for using these probes are less suitable for analysing specific populations of proteins in their native environment. In this study, we address this gap by developing a unimolecular fluorescent probe that combines the properties of small-molecule-based probes and cross-reactive sensor arrays (the so-called chemical 'noses/tongues'). On the one hand, the probe can detect different proteins by generating unique identification (ID) patterns, akin to cross-reactive arrays. On the other hand, its unimolecular scaffold and selective binding enable this ID-generating probe to identify combinations of specific protein families within complex mixtures and to discriminate among isoforms in living cells, where macroscopic arrays cannot access. The ability to recycle the molecular device and use it to track several binding interactions simultaneously further demonstrates how this approach could expand the fluorescent toolbox currently used to detect and image proteins.

Optical cross-reactive sensor arrays (the so-called chemical "noses/tongues") have recently been demonstrated as a powerful tool for high-throughput protein detecting and analysis. Nevertheless, applying this technology to biomarker detection is complicated by the difficulty of non-selective sensors to operate in biological mixtures. Herein we demonstrate a step toward circumventing this limitation by using self-assembled fluorescent receptors consisting of two distinct recognition motifs: specific and non-specific. When combined in an array, binding cooperatively between the specific and non-specific protein binders enables the system to discriminate among closely related isoform biomarkers even in the presence of serum proteins or within human urine.

A method for generating targeted, pattern-generating, protein surface sensors via the self-assembly of modified oligodeoxynucleotides (ODNs) is described. The simplicity by which these systems can be created enabled the development of a sensor that can straightforwardly discriminate between distinct glycoform populations. By using this sensor to identify glycosylation states of a therapeutic protein, we demonstrate the diagnostic potential of this approach as well as the feasibility of integrating a wealth of supramolecular receptors and sensors into higher-order molecular analytical devices with advanced properties. For example, the facile device integration was used to attach the well-known anthracene-boronic acid (An-BA) probe to a biomimetic DNA scaffold and consequently, to use the unique photophysical properties of An-BA to improve glycoform differentiation. In addition, the noncovalent assembly enabled us to modify the sensor with a trinitrilotriacetic acid (tri-NTA)-Ni2+ complex, which endows it with selectivity toward a hexa-histidine tag (His-tag). The selective responses of the system to diverse His-tag-labeled proteins further demonstrate the potential applicability of such sensors and validate the mechanism underlying their function.

Optical cross-reactive sensor arrays (the so-called chemical "noses/tongues") have recently been demonstrated as a powerful tool for high-throughput protein detecting and analysis. Nevertheless, applying this technology to biomarker detection is complicated by the difficulty of non-selective sensors to operate in biological mixtures. Herein we demonstrate a step toward circumventing this limitation by using selfassembled fluorescent receptors consisting of two distinct recognition motifs: specific and non-specific. When combined in an array, binding cooperatively between the specific and non-specific protein binders enables the system to discriminate among closely related isoform biomarkers even in the presence of serum proteins or within human urine.In recent years, extensive efforts have been made to develop non-reductionist approaches to disease diagnosis. Profiling the expression of multiple proteins, rather than detecting individual protein analytes, has been explored as a means of improving diagnostic accuracy and better understanding the parameters affecting disease states.[1] A promising method for obtaining multiplexed protein analysis involves the use of antibodies that can bind and detect the proteins of interest with high affinity and selectivity. [1b,c] This approach has found widespread applications in medical diagnosis; however, the need for producing an antibody for each target and for using stepwise protocols, in addition to their relatively high costs and instability, hamper high-throughput analysis.Cross-reactive sensor arrays, inspired by the mammalian olfactory system, have recently emerged as an alternative detection method that may address these limitations. [2, 3] When the "nose/tongue" approach is used, proteins can be rapidly differentiated using an array of non-specific synthetic receptors that, in combination, generate a unique optical "fingerprint" upon interacting with each protein. Unlike antibody arrays, which operate according to the "lock and key" paradigm, arrays that rely on differential sensing [4] do not require manufacturing multiple antibodies or using technically challenging procedures. As a result, such systems can straightforwardly discriminate among multiple different proteins [3] as well as profile protein mixtures in biofluids, which may indicate disease states. [3f-i] Despite the numerous advantages of cross-reactive arrays, applications for this technology in medical diagnostics are limited by the difficulty of non-selective receptors to operate within biological mixtures. Human serum contains more than 20 000 proteins, of which only about 20 proteins constitute about 99 % of the serum protein mass. Thus, although such systems can effectively discriminate among combinations and concentrations of common serum proteins, [3g] detecting lowabundance disease biomarkers remains challenging. Herein, we present an integrated sensing scheme that uses both the "lock and key" and "differential sensing" strategies for discriminating among low-concentration pr...

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