09/02/2024, 17:44 Chitosan modified graphene field-effect transistor biosensor for ultrasensitive procalcitonin detection - ScienceDirect

Talanta
Volume 268, Part 1, 1 February 2024, 125308

Chitosan modified graphene field-effect transistor

biosensor for ultrasensitive procalcitonin detection
Furong Chen a, Ying Zhang a, Mingxuan Wang a, Jinghai Liu a, Wenfeng Hai a , Yushuang Liu a b

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Abstract

Sepsis is a systemic inflammatory response caused by a bacterial infection that often leading to tissue
damage, organ failure and death. Procalcitonin (PCT), as a peptide precursor to hormones, is the main
biomarker to identification of the sepsis. In this study, a chitosan modified graphene field transistor (CTS-
GFET) was established and first time used for PCT ultra-sensitive detection. CTS was functionalized on the
GFET channel surface to immobilized anti-PCT by glutaraldehyde. This biosensor exhibited the detection
limit as low as 0.82 ag/mL in PBS, which exhibited 3 times enhancement compared with GFET biosensors.
The enhancement mechanisms of CTS-GFET were studied by electrical theoretical model. In addition, the
CTS-GFET biosensor was successfully applied to quantify the concentration of the PCT in human serum
samples, indicating the potential use in clinical application.

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09/02/2024, 17:44 Chitosan modified graphene field-effect transistor biosensor for ultrasensitive procalcitonin detection - ScienceDirect

Introduction

Sepsis, a systemic inflammatory response syndrome caused by bacterial infection, has been threatening
human life, health and safety due to its high infection rate and mortality [[1], [2], [3]]. Although great
progress has been made in anti-infection therapy and organ function support technology, the mortality rate
of sepsis is still as high as 30 %–70 %. Therefore, it is of great significance for the early and sensitive diagnosis
of sepsis to prevent the onset of the septic shock and to increase the survival rate [4,5].

A series of biomarkers have been developed to determinate of sepsis, such as C-reaction protein(CRP),
interleukin 6 (IL-6) and procalcitonin(PCT) [6,7]. Compared with them, PCT can be more specific indicator of
bacterial infection or infection severity. In general, under normal physiological conditions, the PCT level is
below 0.05 ng/mL, while the concentration will be increased at least 1000 folds in serum after the patients
with sepsis within 6–12 h [[8], [9], [10]]. Therefore, a variety of methods such as enzyme–linked
immunosorbent (ELISA) [11,12], fluorescence immunoassay [13], electrochemiluminescence immunoassay
[14,15] and colorimetric immunoassay [16,17] have been developed and used for the qualitative and
quantitative detection of PCT. However, those methods are limited by long analysis times, high costs, and
the need for trained personnel and sophisticated instruments.

Biosensors have been widely used in the field of biochemical molecules and detection because of their
advantages of sensitivity, speed, simple operation and ease of automation. Among the various biosensing
platforms, the graphene-based field-effect transistors (GFETs) are ideal for creating simple and reliable
biosensors due to their fast, sensitive, specific, and low-cost detection properties [18]. Graphene, a single
layer carbon based material, due to its large surface area and good electrical property [19,20], making it an
ideal sensing material for biomolecule. Field effect transistors (FETs) is a three-terminal transistor that uses
an electric field to control the flow of current [21]. For the GFET sensing performance, the specificity and the
action were fully depending on the coupling of effective identification components on the sensing channel
surface. Among the many coupling methods, noncovalent interactions, due to it does not damage the
graphene lattice or reduce its electronic performance, has become the most important coupling method in
GFET sensing. Some bifunctional noncovalent linker, such as 1-pyrenebutanoic acid succinimidyl ester
(PBASE), N-hydroxysuccinimide (NHS) ester tripod, pyrene butyric acid, bovine serum albumin (BSA) has
been successfully used to construct a bio-interface of GFET for the detection of single-point mutations (SNP)
[22]. Exosomes [23]. biotin [24] etc.

Chitosan (CTS), a non-toxic and biocompatible amine rich linear polysaccharide, is synthesized by
diacylation of chitin [25,26]. In general, CTS can be dissolved in an acidic medium and forming layer
structure. The produced film owns high permeability and mechanical strength, good adhesion and
susceptibility to chemical modification [27]. Otherwise, the proton and the counter ions will adjacent to the
CTS chain and create a high-performing dielectrics, which will improve the conductivity and sensitivity for
biological analyses, so CTS has been widely used to fabricate a variety of electronic devices such as
electrochemical sensors, synaptic devices and organic thin film transistors (OTFTs).

Herein, a new method based on non-covalently modified, label-free CTS modified GFET (CTS-GFET)
immunosensor was first time to established and used for PCT quantitative analysis. Using silicon/silicon
dioxide (Si/SiO2) patterned with source and drain electrode as the supporting substrate, graphene was
transferred to the substrate surface as the sensing channel to form the GFET. The CTS was selected as the
linker and adsorbed on the sensing channel surface by the electrostatic adsorption. Anti-procalcitonin
antibodies (anti-PCT) as probe was modified on the CTS surface by glutaraldehyde crosslink to specifically
capture PCT. This is the first report on PCT testing based on CTS-GFET biosensor, which achieve a detection
limit of 0.82 ag/mL and 3 times higher than GFET. The performance enhancement mechanism of CTS-GFET

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09/02/2024, 17:44 Chitosan modified graphene field-effect transistor biosensor for ultrasensitive procalcitonin detection - ScienceDirect

was studied by electrical sensing model and experimental results. The detection of spike serum samples
demonstrated their practical application capabilities.

Section snippets

Materials

PBS buffer and BSA were bought from Solarbio life sciences (Beijing Solarbio Science & Technology Co.
Beijing China). PCT, Anti-PCT, Interleukin-6 (IL-6), C-reactive protein (CRP) were purchased from Cusabio
Biotech CO. (Wuhan China). Chitosan, phosphate buffered solution(PBST) and glutaraldehyde was purchased
from Tansloo (Shanghai, China). 1-pyrenebutanoic acid succinimidyl ester (PBASE), ethanolaine (EA) and
silver were purchased from Sigma-Aldrich. Monolayer graphene was grown by CVD.…

Characterizations

Characterization of CTS-GFET

The morphologies of CTS-GFET biosensor was characterized by SEM. Fig. S2A demonstrated the single layer
graphene transferred on to Si/SiO2 substrate, which present a smooth surface. After chitosan modified, the
graphene surface formed the layered structure and generating some cavities (Fig. S1B). when the anti-PCT
linked on the chitosan surface by the glutaraldehyde, the SEM image showing particulate matter scattered
on the surface of the sensing channel after chitosan modification (Fig. S1C).…

Conclusions

An ultra-sensitive biosensor was developed based on CTS modified GFET for selective detection of PCT. This
CTS-GFET biosensor provide wider detection range (1ag/mL-10 pg/mL) and a LOD as low as 0.82 ag/mL in
PBS buffer, which is more than 3 times sensitivity enhancement compared with that of GFET biosensors.
The enhancement mechanisms of the CTS-GFET biosensor were proposed based on the theoretical model
and experimental results. Additionally, the detection of spiked PCT sample in human serum…

CRediT authorship contribution statement

Furong Chen: Formal analysis, Data curation. Ying Zhang: Formal analysis. Mingxuan Wang: Resources.
Jinghai Liu: Investigation. Wenfeng Hai: Methodology, Funding acquisition. Yushuang Liu:
Conceptualization, Data curation, Writing – review & editing, Project administration.…

Declaration of generative AI in scientific writing

The authors declare not to use the generative artificial intelligence (AI) and AI-assisted technologies to
analyses and draw insights from data as part of the research process, even in the writing process.…

Declaration of competing interest

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We declare that we have no financial and personal relationships with other people or organizations that can
inappropriately influence our work, there is no professional or other personal interest of any nature or kind
in any product, service and company that could be construed as influencing the position presented in, or the
review of, the manuscript entitled.…

Acknowledgements
This work was supported by the National Natural Science Foundation of China (22104066, 22164016).
Innovation and entrepreneurship start-up project for students studying in Inner Mongolia (2021LXCX002).
Inner Mongolia Natural Science Foundation (2020BS08007, 2020BS02007). Program for Young Talents of
Science and Technology in Universities of Inner Mongolia Autonomous Region (NJYT23034) and Ph. D Start-
up Fund of Inner Mongolia Minzu University (BS515, BS453).…

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