Cucurbit[n]uril-Immobilized Sensor Arrays for Indicator-Displacement Assays of Small Bioactive Metabolites

Abstract

The patterned immobilization of chemosensors into nano/microarrays has often boosted utilization in diagnostics and environmental sensing applications. While this is a standard approach for biosensors, e.g., with antibodies, other proteins, and DNA, arraying is not yet adopted widely for supramolecular chemosensors which are still predominantly used in solution systems. Here we introduce the patterned immobilization of cucurbit[n]urils (CBn) into multiplexed microarrays and elucidate their prospects for the advancement of surface-bound indicator-displacement assays to detect small molecule analytes. The microarrays were generated by microchannel cantilever spotting of functionalized CBn and subsequent self-assembly of the corresponding indicator dyes from solution. Enhanced sensitivity of surface-bound microarrays was established in demonstrations with small bioactive metabolites (spermine, amantadine, and cadaverine) compared to bulk assays. Furthermore, the integration of the CBn/indicator microarrays into microfluidic channels provides an efficient route for real-time monitoring of the sensing process, allows easier handling, and reduces need for analyte volume. The concept was further extended to differential sensing of analytes on diplex or multiplex CBn/indicator microarrays, opening up a route for multicomponent sensing of small molecule analytes in complex liquids.

A molecular-engineered rotaxane overcomes longstanding biofluid-challenges and enables optical tryptophan detection in human serum

“…The remarkable sensitivity increase of rotaxane 1 by surface immobilization corresponds to our recent findings for indicator displacement chemosensors. 60 Importantly, the rotaxane-functionalized microarrays are more selective than printed microarrays with the binary CB8•MDAP chemosensor complex. While both allow for detection of tryptophan or indole, the hydrophobic and strongly CB8-binding guest memantine does not alter the emission of the rotaxane 1 microarray.…”

“…By encapsulation of such sensor chips into microfluidic channels, they can act as convenient and low volume sensing platform. 60 The possibility to immobilize the Trp-binding rotaxane on surfaces opens up its potential for the future integration into lab-on-achip designs with plasmonic biosensors. 62…”

“…After printing, the substrate was heated to 80°C for 24 h, then washed with ethanol, and dried with N2. Control experiments were performed according to previous work, 60 see also Supplementary Information.…”

“Writing biochips”: high-resolution droplet-to-droplet manufacturing of analytical platforms

“…Some relevant applications of in the field of bioanalysis have recently been demonstrated by the same research group of Hirtz, in handling different kinds of biomolecular inks ( e.g. peptides, 104 phospholipids, 105 proteins, 106 small molecules 107 ), resulting in the realization of microarrays for bioanalysis with optical detection.…”

“…In another recent publication, they realized glycoluril-based cucurbit[ n ]uril (CBn) microrarray sensors by displacement assays of fluorescent indicators (for example tetramethylrhodamine cadaverin) for different molecular systems (spermine, cadaverine and amantadine), reaching limits of detection down to nanomolar concentration ranges. 107 Finally, a remarkable achievement was also the realization of macrophage capture microarrays onto surface-modified glass supports. 112 To this aim, the authors used poly(oligoethylene glycol methylether methacrylate) functionalized to expose azide groups which in turn permitted the click chemistry functionalization via μCS of biotin–dibenzocyclooctyne spots.…”