bioRxiv Science⌕ Search
Home/Archive/bioRxiv

bioRxiv · 10.1101/273078

Femtomolar and selective dopamine detection by a gold nanoparticle enhanced surface plasmon resonance aptasensor

Cao, Y.·McDermott, M. T.
View original source ↗Export metadata ↓

Abstract

Ultrasensitive and selective detection and quantification of dopamine (DA) plays a key role in monitoring neurodegenerative diseases. However, the detection limit reported for DA detection is typically in the lower nM range. Pushing the detection limit to pM or lower for this particular target to cover the physiological levels (< 130 pM) is significant. Herein, DA DNA aptamer (DAAPT) gold nanoparticle (AuNP) conjugate is utilized to enhance the surface plasmon resonance (SPR) signal, which enables to detect and quantify DA in the femtomolar (200 fM) to picomolar range. To the best of our knowledge, this is the lowest detection limit achieved for SPR sensing of dopamine. The as-prepared 10 nm DAAPT-AuNP conjugate demonstrates strong binding affinity (Kd = 3.1 {+/-} 1.4 nM) to the complementary DNA (cDNA) probe on gold chip. The cDNA probe is immobilized to the chip via polydopamine surface chemistry, which allows the Michael addition of any primary amine-terminated biomolecules. By adjusting the concentration of the DAAPT-AuNP conjugate, two calibration curves are generated with dynamic ranges from 100 {micro}M to 2 mM, and from 200 fM to 20 nM, respectively. Both calibration curves have negative slopes, showing good agreement to a dose-response curve in an enzyme inhibition assay. In addition, the sensing strategy is evaluated to be specific for DA detection using a series of DA analogs and other metabolites as potential interferences.

Explore related subjects

biochemistryPreprint

Keep this discovery

Explore connections, maps & timelines

BibTeXRIS

Cao, Y., McDermott, M. T.. 2018-02-28. Femtomolar and selective dopamine detection by a gold nanoparticle enhanced surface plasmon resonance aptasensor. https://doi.org/10.1101/273078

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related preprints

bioRxiv2026

Functional Reconstitution of Human OATP1B1 and OATP1B3 in Proteoliposomes: Effect of Bicarbonate Gradients and Substrate-specific Stimulation on Transport

The hepatic organic anion transporting polypeptides OATP1B1 and OATP1B3 are critical for the clearance of widely prescribed drugs, hepatotoxins, and endogenous metabolites. However, the thermodynamic driving force governing their transport has remained a subject of debate due to the complexity of cellular models. Here, we report a defined, reconstituted proteoliposome system to study the transport kinetics of isolated human OATP1B1 and OATP1B3. Through direct functional assays, real-time counter-ion tracking, and a novel ABCG2-OATP1B1 functional co-reconstitution, we identify an outwardly directed bicarbonate gradient as a thermodynamic driving force for concentrative substrate uptake. In contrast we find no evidence for the proposed role of reduced glutathione efflux as a possible driving force. Targeted mutagenesis experiments revealed that the conserved TM3 residue His115 is needed to couple the bicarbonate gradient to concentrative substrate uptake, mechanistically linking substrate antiport to local pH-sensitive bicarbonate processing. We also investigated the substrate-specific stimulatory effect of the green tea catechin epigallocatechin gallate (EGCG) on OATP1B3 transport, confirming that this effect is intrinsic to OATP1B3, substrate specific, and does not need the involvement of other cellular proteins. Our findings define the energetic basis of OATP1B1 and OATP1B3 hepatic xenobiotic clearance.

biochemistry↗
bioRxiv2026

Mapping the sequence determinants of constitutive activity in GPCRs

G protein-coupled receptors (GPCRs) signal not only in response to ligands but also in their absence. This basal, or constitutive, activity has distinct physiological consequences and is the only functional measurement available for orphan receptors. It is also hard to compare receptors because each is reported relative to its own baseline. Here, we measure basal activity on a single scale across the human GPCR family. Pairing a barcoded library of all human GPCRs with a transcriptional cAMP reporter, we scored 901 receptor constructs in one pool experiment. Basal activity forms a continuous distribution, and many of the highest-scoring recipes have prior reports of ligand-independent activity, with orphan being strongly represented. We then exhaustively mutated one of them, GPR161, and measured its effects on protein abundance, surface expression, and cAMP signaling. Of the 528 positions, only 10 specifically alter signaling independent of expression. Constraints on basal signaling are dominated by reepotro biogenesis, which requires multiphenotype scanning to resolve. Together, these datasets place basal activity on a common scale across the GPCR family and provide a template for dissecting it at residue resolution.

biochemistry↗
bioRxiv2026

Direct Bioconjugations of Diverse Payloads using an Extraordinarily Donor-Promiscuous Flavin Transferase

Site-specific protein conjugation is a widely used tool in chemical biology research and is the foundation of many biotechnological and biomedical applications. A key enabling technology are repurposed post-translational protein modifying enzymes, where variants with broad donor and acceptor substrate promiscuity were developed into general-purpose bioconjugation platforms. However, direct enzymatic attachment of bulky or structurally diverse payloads is often limited by low reaction efficiencies and difficulties in preparing suitable donors. Consequently, conjugation typically relies on two-step strategies in which an enzyme first installs a bioorthogonal handle on the target protein, followed by chemical attachment of the payload. These enzyme-first, chemistry-second approaches limit chemistry to bioorthogonal reactions that leave a molecular footprint. Here, we introduce direct ADDing, an enzymatic bioconjugation platform based on our discovery of the exceptional donor promiscuity of the flavin transferase ApbE. We show that ApbE accepts a remarkably broad range of donor substrates provided they carry an adenosine diphosphate (ADP) anchor. The prevalence of ADP-containing compounds in nature provides access to diverse building blocks and an extensive biochemical toolbox for donor generation. Exploiting these resources, we demonstrate multiple strategies for preparing compatible donors and achieve direct, site-specific enzymatic conjugation of diverse small molecules, peptides, proteins, and synthetic polymers.

biochemistry↗