Label-free detection of glycoproteins by the lectin biosensor down to attomolar level using gold nanoparticles

Abstract

We present here an ultrasensitive electrochemical biosensor based on a lectin biorecognition capable to detect concentrations of glycoproteins down to attomolar (aM) level by investigation of changes in the charge transfer resistance (Rct) using electrochemical impedance spectroscopy (EIS). On polycrystalline gold modified by an aminoalkanethiol linker layer, gold nanoparticles were attached. A Sambucus nigra agglutinin was covalently immobilised on a mixed self-assembled monolayer formed on gold nanoparticles and finally, the biosensor surface was blocked by poly(vinyl alcohol). The lectin biosensor was applied for detection of sialic acid containing glycoproteins fetuin and asialofetuin. Building of a biosensing interface was carefully characterised by a broad range of techniques such as electrochemistry, EIS, atomic force microscopy, scanning electron microscopy and surface plasmon resonance with the best performance of the biosensor achieved by application of HS-(CH2)11-NH2 linker and gold nanoparticles with a diameter of 20 nm. The lectin biosensor responded to an addition of fetuin (8.7% of sialic acid) with sensitivity of (338 ± 11) Ω decade(-1) and to asialofetuin (≤ 0.5% of sialic acid) with sensitivity of (109 ± 10) Ω decade(-1) with a blank experiment with oxidised asialofetuin (without recognisable sialic acid) revealing sensitivity of detection of (79 ± 13) Ω decade(-1). These results suggest the lectin biosensor responded to changes in the glycan amount in a quantitative way with a successful validation by a lectin microarray. Such a biosensor device has a great potential to be employed in early biomedical diagnostics of diseases such as arthritis or cancer, which are connected to aberrant glycosylation of protein biomarkers in biological fluids.

Electrochemical detection of glycan and protein epitopes of glycoproteins in serum

“…[25][26][27]30 In contrast, a linear increase from baseline impedance was observed when the SNA lectin biosensor was incubated with an increasing concentration of fetuin expressing a correct glycan epitope. [25][26][27]30 Apart from showing very high binding specicity towards the glycan epitope, these SNA lectin biosensors have demonstrated sensitivity for glycoprotein detection (e.g. fetuin) up to attomolar concentration.…”

“…29 In fact, glycan structure specic interactions of a glycoprotein with lectins are very well demonstrated in the recent past mainly using sialic acid binding lectin biosensors. [25][26][27]30 Incubation of the SNA lectin biosensor with asialofetuin, a non-sialic acid expressing glycoform variant of glycoprotein fetuin, showed very minimal change in the baseline impedance. [25][26][27]30 In contrast, a linear increase from baseline impedance was observed when the SNA lectin biosensor was incubated with an increasing concentration of fetuin expressing a correct glycan epitope.…”

“…[25][26][27]30 Incubation of the SNA lectin biosensor with asialofetuin, a non-sialic acid expressing glycoform variant of glycoprotein fetuin, showed very minimal change in the baseline impedance. [25][26][27]30 In contrast, a linear increase from baseline impedance was observed when the SNA lectin biosensor was incubated with an increasing concentration of fetuin expressing a correct glycan epitope. [25][26][27]30 Apart from showing very high binding specicity towards the glycan epitope, these SNA lectin biosensors have demonstrated sensitivity for glycoprotein detection (e.g.…”

Glycoprofiling as a novel tool in serological assays of systemic sclerosis: A comparative study with three bioanalytical methods

“…1) and EIS-based lectin biosensor down to fM-aM level [28,32,52]. From all these three methods of glycoprofiling lectin microarray is the best, followed by ELLA and EIS-based lectin biosensors in terms of multiplexing, while in case sensitivity of assays is the main requirement than this order is completely reversed (Tab.…”

Perspectives in Glycomics and Lectin Engineering

“…In our recent work we focused on the development of ultrasensitive impedimetric lectin biosensors with detection limits down to a single-molecule level based on controlled architecture at the nanoscale (59)(60)(61). In the first study the biosensor was able to detect a glycoprotein in a concentration window spanning 7 orders of magnitude with a detection limit for the glycoprotein down to 0.3 fM, what was the lowest glycoprotein concentration detected (59).…”