Arabian Journal of Chemistry (2023) 16, 104582

King Saud University

Arabian Journal of Chemistry

www.ksu.edu.sa
www.sciencedirect.com

ORIGINAL ARTICLE

ssDNA-QDs/GO multicolor fluorescence system

for synchronous screening of hepatitis virus DNA
Junping Guo 1, Huan Zhang 1, Jiajia Yang *, Ying Zhang, Jiekai Wang, Guiqin Yan

Shanxi Normal University, Taiyuan 030031, China

Received 23 August 2022; accepted 10 January 2023

Available online 16 January 2023

KEYWORDS Abstract Viral hepatitis is a common infectious disease caused by five viruses (hepatitis virus A, B,
Multicolor quantum dot; C, D, and E). Given the diversity of hepatitis virus, rapid screening and accurate typing of viral hep-
Synchronous screening; atitis are the prerequisites for hepatitis therapy. Here, a multicolor fluorescence system was con-
Graphene oxide; structed by combining with the multi-color fluorescence properties of CdSe/ZnS quantum dots
Hepatitis virus DNA (QDs, emission wavelengths: 525 nm, 585 nm and 632 nm) and the broad-spectrum fluorescence
quenching performance of GO. Taking advantage of the specific recognition of ssDNA modified
CdSe/ZnS QDs to target hepatitis virus DNA, the constructed system could effectively distinguish
hepatitis A virus DNA (HAV-DNA), hepatitis B virus DNA (HBV-DNA), and hepatitis C virus
DNA (HCV-DNA) in a homogeneous solution. Based on the different adsorption property of
GO for ssDNA and dsDNA, the fluorescence Forster resonance energy transfer (FRET) process
between ssDNA modified QDs and GO could be regulated. The fluorescence signal of the con-
structed system presented a sensitive response to HAV-DNA, HBV-DNA, and HCV-DNA content
in the range of 1.0–192 nM, 8.0–192 nM, and 1.0–128 nM, respectively. The limit of detection for
HAV-DNA, HBV-DNA, and HCV-DNA is 0.46 nM, 1.53 nM, and 0.58 nM. The constructed sys-
tem can be used to screen hepatitis virus DNA in real samples, which provides an alternative strat-
egy for rapid screening and diagnosis of viral hepatitis.
Ó 2023 The Author(s). Published by Elsevier B.V. on behalf of King Saud University. This is an open
access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

1. Introduction

Hepatitis defined as the inflammation of the liver tissue is a globally

common disease. Although there are many pathogenic factors that
* Corresponding author. can induce hepatitis (viruses, bacteria, parasites, chemical poisons,
E-mail address: [email protected] (J. Yang). drugs, alcohol, and autoimmune factors), viral hepatitis remains the
1
These authors contributed equally and share first name co- predominant cause for liver disease and has been a global public health
authorship. problem due to its strong contagiousness, complex transmission route
Peer review under responsibility of King Saud University. and high incidence (Semaine et al., 2006; van Leeuwen et al., 2022).
According to statistics, approximately 1.3 million people die as a result
of viral hepatitis each year worldwide, which is mainly attributed to
cirrhosis and liver cancer caused by hepatitis B virus (HBV), hepatitis
Production and hosting by Elsevier C virus (HCV), hepatitis D virus (HDV), acute infection of hepatitis A

https://doi.org/10.1016/j.arabjc.2023.104582
1878-5352 Ó 2023 The Author(s). Published by Elsevier B.V. on behalf of King Saud University.
This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
2 J. Guo et al.

virus (HAV) and liver failure caused by hepatitis E virus (HEV) When QDs and GO are close to each other, the energy will transfer
(Krugman et al., 1997). Given the diversity of viral hepatitis types from the QDs to the GO and then the fluorescence signal of QDs will
and the limited cross-immunization between them, humans can be be quenched (Zhang et al., 2011; Arvand & Mirroshandel, 2019).
co-infected, or overlapping infected with hepatitis viruses. Thus, diag- Therefore, promoting or preventing the interaction between QDs
nosis and classification of viral hepatitis is still a challenging topic. and GO by the mediation of targets has been used as an ingenious
Viral hepatitis can be diagnosed using serology tests (antibody) or strategy for the construction of various probes (Arvand &
molecular (presence of viral RNA or DNA) diagnostics. Hepatitis Mirroshandel, 2019). Numerous studies have demonstrated that GO
virus DNA detection is an effective method with high assay efficiency can strongly adsorb single-stranded DNA (ssDNA) via hydrophobic
and low sample requirement. Up to now, a large number of DNA and p-p stacking interactions between DNA nucleobases and sp2-
detection methods have been developed for the diagnosis of viral hep- hybridized atoms of GO nanosheets, but it has low affinity toward
atitis including high performance liquid chromatography(Wolford double-stranded DNA (dsDNA)(Liu et al., 2016a; Wang et al.,
et al., 2000), real-time polymerase chain reaction (PCR) (Geng et al., 2018). Based on these unique properties of GO (Park et al., 2013;
2006; Wang et al., 2013b), surface-enhanced Raman scattering(Zhu Wu et al., 2011), several ssDNA modified QDs/GO fluorescent probes
et al., 2021), electrochemical detection(Manzano et al., 2018; Wang have been reported (Arvand & Mirroshandel, 2019; Li et al., 2017).
et al., 2013a; Wang et al., 2013b), and colorimetric detection(Mao These studies provided valuable references for accurate detection tar-
et al., 2016), molecularly imprinted polymers-RLS sensor, resonance gets in complex matrices. However, they mainly focus on the detection
light scattering sensor, molecular imprinting fluorescence sensor of single target. Combined with our previous work (Yang et al., 2018)
(Luo, et al., 2016), gas-responsive molecularly imprinted sensor. These and inspired by the research of Xiang et al. (Xiang et al., 2020), the
strategies are highly sensitive, but most of them are employed to detect construction of QDs/GO composite fluorescence system may be an
a single type of hepatitis virus DNA. Accordingly, it is difficult to effective approach to realize the synchronous screening of multiple
achieve rapid typing of hepatitis types. Due to the limited information viral hepatitis DNA.
provided by single target analysis, false-positive results are inevitable, In this paper, a multicolor fluorescence system was attempted to
which will result in a relatively high rate of misdiagnosis and missed construct for synchronous screening of hepatitis virus DNA. As illus-
diagnosis (He et al., 2017). In order to overcome these shortcomings, trated in Scheme 1, three water-soluble CdSe/ZnS QDs with different
multiplexed target DNA detection will be an ideal approach for accu- maximum emission wavelength at 525 nm, 585 nm and 632 nm were
rate diagnosis of viral hepatitis. firstly modify by using of complementary ssDNA strands of HAV-
Compared with single detection, multi-biomarkers simultaneous DNA, HBV-DNA, and HCV-DNA though cross-linking reaction.
analyses are becoming increasingly popular, which enable to detect Taking advantage of the multicolor fluorescence property of CdSe/
two or more targets in a single analyze with high detection flux, low ZnS QDs and unique quenching performance of GO, a multicolor flu-
sample requirement, shorter assay time, and low expenditure(Liu orescence response system was constructed, in which QDs would be
et al., 2016b). Currently, several materials have been used for simulta- adsorbed on the surface of GO by the mediation ssDNA ligands,
neous detection, such as organic fluorescence dyes (Ren et al., 2022), and the fluorescence signals of QDs would be synchronously quenched
quantum dots (QDs) (Hu et al., 2021; Jie et al., 2017; Li et al., through FRET. When the HAV-DNA, HBV-DNA, and HCV-DNA
2020), metal nanoparticles (Kim & Jurng, 2011; Zhang et al., 2020), were captured by complementary ssDNA ligands modified QDs
and so on. Although organic fluorescent dyes have been successfully through hybridization reaction, FRET will be blocked due to weak
applied to the simultaneous detection of multiple targets, the simulta- adsorption of GO to double-stranded DNA (dsDNA), and strong flu-
neous detection of multiple targets requires multiple excitation opera- orescence signals will be detected. Compared with single fluorescence
tions as the inconsistent excitation wavelength of each organic and double-color fluorescence detection, a multicolor fluorescence sys-
fluorescent dye restricts them from being excited synchronously at tem was proposed by virtue of QDs and GO, which is expected to serve
the same excitation wavelength. Compared with fluorescent dyes, the as an alternative strategy for rapid screening and diagnosis of viral
multicolor fluorescence properties of QDs under single excitation hepatitis.
wavelength endow them with excellent flexibility in developing syn-
chronous detection system (Chandan et al., 2018; Shen et al., 2021; 2. Experimental section
Xiang et al., 2020). Especially, QDs can be assembled with a variety
of nano-materials to form composite detection systems (Adegoke &
Forbes, 2016; Miao et al., 2016). Typically, QDs/GO composite system
2.1. Materials and chemicals
can be served as an excellent platform for target detection based on flu-
orescence resonance energy transfer (FRET), wherein QDs is used as The DNA sequences (shown in Table S1) used in the experi-
energy donor and GO acts as energy acceptor (Zhang et al., 2011). ment were synthesized and purified by Sangon Biotechnology

Scheme 1 Schematic illustration of ssDNA-QDs/GO multicolor fluorescence system for synchronous screening of hepatitis virus DNA.
ssDNA-QDs/GO multicolor fluorescence system 3

Co. ltd. (Shanghai, China). N-Hydroxylsuccinimide sodium of 340 nm, and the slit widths for excitation and emission were
salt (NHS) and 1-ethyl-3-(3-(dimethylamino) propyl) carbodi- set at 10 nm.
imide hydrochloride (EDC) were purchased from Sigma-
Aldrich (St. Louis, USA). Three PEG-capped and carboxyl- 2.5. Screening of hepatitis virus DNA in real sample
modified CdSe/ZnS QDs (the maximum emission wavelength
at 525 nm, 585 nm and 632 nm, the excitation wavelength at The serum samples were purchased from Sigma-Aldrich Com-
340 nm) were provided by Suzhou Xingshuo Nanotech Co., pany, which were diluted 10 times with distilled water before
ltd. (Suzhou, China). GO was purchased from Nanjing Xian- use. Then,16 nM and 64 nM of the target hepatitis virus
feng Nanomaterials Technology Co., ltd. (Nanjing, China). DNA were added to the samples, respectively. The labeled
The other reagents used were of analytical grade, and all the samples with different standard additions were examined at
solutions were prepared with twice-distilled water. an excitation wavelength of 340 nm under the optimized con-
ditions, and the fluorescence spectra of the QDs@CDNA and
2.2. Instruments GO/DNA system were observed in the range of 400–700 nm.

The morphologies of all samples were characterized by high- 3. Results and discussion
resolution transmission electron microscope (HR-TEM)
(JEM-2100, Hitach, Japan). Fourier transform infrared spec- 3.1. Construction of multicolor fluorescence system
troscopy was performed on a Magna-560 spectrometer (Nico-
let, Madison, WI). The UV–vis absorption spectra were
Three kinds of QDs have good dispersion and spherical mor-
monitored using a Shimadzu UV-29100 spectrophotometer
phology (Fig. S1A-C). The average hydrodynamic size of
(Tokyo, Japan). All fluorescence spectra were measured on a
QDs(5 2 5), QDs(5 8 5), and QDs(6 3 2) is 11.62 nm, 12.54 nm,
Cary Eclipse fluorescence spectrophotometer (Varian Ameri-
and 13.44 nm, respectively (Fig. S1D-F). In order to verify
can Pty ltd., USA) with a 1 cm path-length quartz cell. The
the feasibility of constructing multicolor fluorescence system
excitation wavelength was 340 nm, and the silt widths of exci-
using QDs with different maximum emission wavelength, the
tation and emission were set at 10 nm.
fluorescence emission spectrums of QDs(5 2 5), QDs(5 8 5),
and QDs(6 3 2) were firstly measured with the same concentra-
2.3. Preparation of ssDNA modified QDs
tion at the exciting wavelength of 340 nm. As presented in
Fig. 1A, the maximum emission intensities of QDs(5 2 5),
Three ssDNA modified QDs were prepared as follows: 23.0 lL QDs(5 8 5), and QDs(6 3 2) corresponded to 525 nm, 585 nm,
EDC (10 mg mL 1) and 40 lL QDs (10 lmol/L) were added to and 632 nm, respectively. It is consistent with the instruction
200 lL PBS buffer (100 mmol/L, pH = 6.0). The solution was manual provided by Suzhou Xingshuo Nanotech Co., ltd,
continuously and gently agitated for 30 min to activate the car- and can be found on the company website. Besides, it also
boxyl group. Subsequently, the pH of the above solution was could be found that there is no obvious overlap between the
adjusted immediately to 8.0 with PBS (pH = 8.5) prior to emission spectrum of three QDs, which indicated that the
the addition of 13.8 mL NHS (10 mg mL 1) and a certain con- emission signal of each QDs can be detected individually in a
centration of complementary DNA. The mole ratio of comple- mixed system of QDs(5 2 5), QDs(5 8 5), and QDs(6 3 2). More-
mentary DNA to QDs was 40:1. After gently shaking for 3 h at over, it was exciting that no significant change was observed in
room temperature, the resulting solutions were placed in a the fluorescence intensity of QDs(5 2 5) after QDs(5 8 5) and
refrigerator at 4 ℃ overnight. Then, an ultrafilter tube (30 QDs(6 3 2) were added in succession, and the emission spec-
kD) was used to centrifuge the solution to remove excess trum of QDs(5 2 5) did not show obvious blue-shift and red-
ssDNA, EDC, and NHS. The final products were diluted to shift (Fig. 1B). The similar results were also observed even if
5 mL for the subsequent experiments. the sampling sequence of the QDs was swapped (Fig. 1C, D).
In order to realize the selective recognition of three hepatitis
2.4. Synchronous screening of hepatitis virus DNA viral DNA, QDs(5 2 5), QDs(5 8 5), and QDs(6 3 2) was modi-
fied by complementary ssDNA strands of HAV-DNA, HBV-
The as-prepared QDs@CHAV-DNA, QDs@CHBV-DNA, and DNA and HCV-DNA, respectively. The modified QDs were
QDs@CHCV-DNA solution with the same concentration was referred to as QDs@CHAV-DNA, QDs@CHBV-DNA, and
mixed together in equal proportions. The mixed solution was QDs@CHCV-DNA. Compared with the unmodified QDs, typi-
abbreviated as QDs@CDNA. Then, 60 mL QDs@CDNA mixed cal DNA absorption peaks between 260 and 280 nm can be
solution and 500 mL Tris-HCL buffer (20 mmol/L Tris-HCL, observed in UV–vis absorption spectrums of the QDs@C-
0.1 mol/L NaCl, pH = 7.4) were added to a series of 5.0 mL col- HAV-DNA, QDs@CHBV-DNA, and QDs@CHCV-DNA (Fig. S2),
orimeter tubes. And then HAV-DNA, HBV-DNA and HCV- indicating the ssDNA strands were successfully modified on
DNA solution with different concentrations were added and a the surface of QDs. As shown in Fig. 2A, the fluorescence
constant volume of 1 mL was achieved using distilled water. intensity of modified QDs did not change obviously relative
After mixed thoroughly and hybridized for 60 min in a ther- to those of unmodified QDs. The solution of QDs@CHAV-
mostat water bath at 37 ℃, 150 mL GO (0.5 mg mL 1) was added DNA, QDs@CHBV-DNA, and QDs@CHCV-DNA presented
into the above mixed solution, and the solution volume was green, yellow and pink fluorescence under UV illumination,
diluted to 2.5 mL with distilled water and incubated for respectively (Fig. 2B). The kinetic analysis of the change of flu-
30 min at room temperature. The fluorescence spectra were orescence intensity of three modified QDs showed that the flu-
recorded from 400 nm to 700 nm at an excitation wavelength orescence intensity of QDs@CHAV-DNA, QDs@CHBV-DNA,
4 J. Guo et al.

Fig. 1 (A)The emission fluorescence spectra of pure QDs(5 2 5), QDs(5 8 5) and QDs(6 3 2). (B)-(D) Comparison of the fluorescence
intensity of a single QDs and that of mixture QDs.

and QDs@CHCV-DNA remained relatively stable within 60 min QDs@CHCV was approximately 79.71 %, 78.04 %, and
(Fig. S3A-C). The fluorescence spectrums of QDs@CDNA 84.56 %, respectively. Thus, the fluorescence of QDs@CHAV,
remained almost unchanged (inset in Fig. S3 A-C). The quan- QDs@CHBV and QDs@CHCV could be quenched simultane-
tum yield of QDs(5 2 5), QDs(5 8 5), and QDs(6 3 2) was ously by GO.
83.4 %, 87.3 %, 89.5 %, respectively. After modification with As shown in Fig. 2D, when HAV-DNA was incubated with
HAV-DNA, HBV-DNA, and HCV-DNA, the quantum yield QDs@CHAV-DNA for 60 min prior to the addition of GO, the
of QDs@CHAV-DNA, QDs@CHBV-DNA, and QDs@CHCV- fluorescence intensity of QDs@CHAV-DNA was obviously
DNA was 80.1 %, 83.1 %, 84.6 %, respectively. It indicated recovered at 525 nm, but there was no obvious change of the
that ssDNA modification does not change the fluorescence fluorescence intensity at 585 nm and 632 nm. If the HBV-
properties of QDs(5 2 5), QDs(5 8 5), and QDs(6 3 2). These DNA or HCV-DNA was incubated with QDs@CHBV-DNA
results indicated that the QDs@CHAV-DNA, QDs@CHBV- or QDs@CHCV-DNA before adding GO, the fluorescence inten-
DNA, and QDs@CHCV-DNA have strong fluorescence proper- sity of the QDs@CHBV-DNA and QDs@CHCV-DNA at 585 nm,
ties and good optical stability. and 632 nm also recovered obviously, illustrating the proposed
As aforementioned, GO can effectively adsorb ssDNA via system can effectively recognize and distinguish three kinds of
hydrophobic and p-p stacking interactions between DNA target DNA(Fig. S5A-B). If the DNAHAV, DNAHBV, and
nucleobases and sp2-hybridized atoms of GO nanosheets, but DNAHCV were mixed in equal proportion and incubated with
it has low affinity toward double-strand DNA (dsDNA) (Liu QDs@CDNA together, the fluorescence intensity was recovered
et al., 2016a; Wang et al., 2018). Besides, GO can effectively simultaneously at 525 nm, 585 nm and 632 nm (Fig. S5C). This
quench the fluorophore of QDs through energy resonance further confirmed that current conception is feasible and can
transfer. Here, one of our concerns is that whether the fluores- be used for constructing a synchronous screening system for
cence of QDs with three emission wavelengths can be syn- three kinds of hepatitis virus DNA.
chronously quenched. Consequently, the fluorescence
quenching properties of GO for QDs@CHAV-DNA, QDs@C- 3.2. Optimization of conditions for synchronous screening of
HBV-DNA, and QDs@CHCV-DNA were investigated. From hepatitis viral DNA
Fig. S4 it could be seen that the GO is transparent due its sin-
gle layered structure and has irregular edges. As illustrated in The effect of GO concentration on the fluorescence intensity of
Fig. 2C, the fluorescence intensity of the mixed system signifi- the multicolor fluorescence system was investigated. As shown
cantly reduced after the addition of GO, and the fluorescence in Fig. S6A, the fluorescence intensity corresponding to
quenching efficiency for QDs@CHAV, QDs@CHBV and 525 nm, 585 nm and 632 nm decreased gradually with the
ssDNA-QDs/GO multicolor fluorescence system 5

Fig. 2 (A)The emission fluorescence spectra of the modified QDs and the unmodified QDs. (B) Photograph of the QDs@CHAV-DNA,
QDs@CHBV-DNA, and QDs@CHCV-DNA under UV illumination. (C) The change of fluorescence emission spectra of QDs@CDNA in
presence of GO (30 lg mL 1). (D) Fluorescence emission spectra of QDs@CDNA/GO with different concentrations of HAV-DNA
(1.0 nM and 128 nM).

increase of GO concentration. From the response curve of the could be seen that the fluorescence intensity did not change
fluorescence intensity ratio (F/F0) to GO concentration (where when the GO concentration was higher than 30 lg mL 1
F and F0 refer to the fluorescence intensity of the multicolor (Fig. S6 B). Thus, 30 lg mL 1 GO was selected for the subse-
system in the presence and absence of GO, respectively), it quent experiments.

Fig. 3 (A) Fluorescence spectra of the multicolor fluorescence system with different concentrations of HAV-DNA, HBV-DNA, and
HCV-DNA (0, 1.0, 8.0, 16.0, 32.0, 64.0, 128.0, 192.0 nM). (B)The relationships between fluorescence intensity ratio (F/F0) and the
concentration of HAV-DNA, HBV-DNA, and HCV-DNA.
6 J. Guo et al.

As we know, weak alkaline medium is favorable for DNA target DNA, respectively) and the concentration of three hep-
hybridization. The effect of pH on fluorescence intensity of the atitis virus DNA. The F/F0 presented a sensitive response to
multicolor fluorescence system was investigated over the pH the concentration of HAV-DNA, HBV-DNA, and HCV-
range of 6.0–8.5 (Fig. S6C). The fluorescence quenching effi- DNA in the range of 1.0–192 nM, 8.0–192 nM, and 1.0–
ciency of GO for three kinds of QDs@CDNA was relatively 128 nM, respectively. The standard curves for HAV-DNA,
high at pH 7.4, but decreased at pH higher than 7.4. In the HBV-DNA and HCV-DNA were F/F0 = 0.012C(HAV-
2
presence of target DNA, the maximum fluorescence recovery DNA) + 1.19 (R = 0.992), F/F0 = 0.0102C(HBV-
2
ratio was achieved at pH = 7.4. In order to ensure the high DNA) + 1.27(R = 0.984) and F/F0 = 0.0132C(HCV-
2
sensitivity of the system, Tris-HCl buffer with pH 7.4 was DNA) + 1.25 (R = 0.985), respectively. Based on 3r/S, the
selected for the subsequent experiments. detection limit for HAV-DNA, HBV-DNA, and HCV-DNA
Fig. S6D displayed the effects of incubation time on analyt- were calculated as 0.46 nM, 1.53 nM, and 0.58 nM. The rela-
ical performance of the multicolor fluorescence system. The tive standard deviation (RSD) for five times repeated detection
fluorescence intensity of the system in the presence of GO of 32 nM HAV-NDA, HBV-DNA, and HCV-DNA was about
gradually decreased with the prolonged incubation time and 3.07 %, 3.73 %, 3.45 %, respectively. It indicated acceptable
plateaued within 30 min at room temperature. Thus, 30 min reproducibility of the proposed multicolor fluorescence system.
was selected as the reaction time between QDs@CDNA and Table 1 shows the comparison between the proposed
GO. The hybridization reaction time of QDs@CDNA/GO method and other reported methods for hepatitis virus DNA
and target DNAs was further optimized. It could be seen that in terms of detection range and limit. The analytical parame-
the fluorescence intensity ratio of the system gradually ters of the constructed system are comparable to or better than
increased with the extension of hybridization time, and did
not obviously change after 60 min (Fig. S6E). Thus, 60 min
was selected as the hybridization time.
The ionic strength played an important role in DNA
hybridization reaction. the fluorescence recovery ratio F/F0
was optimal when the NaCl concentration in hybridized buffer
was 50 mM (Fig. S6F). Therefore, 50 mM NaCl solution was
used in this study.

3.3. Synchronous screening of hepatitis viral DNA

The analytical performance of the constructed system was

evaluated under the optimized conditions by adding three
kinds of hepatitis viral DNA with the same proportion. As
shown in Fig. 3A, the fluorescence intensities of the system
Fig. 4 Selectivity investigation of the proposed method in the
at 525 nm, 585 nm, and 632 nm simultaneously increased with
presence of target DNA and different base mismatch strands. The
the increase of HAV-DNA, HBV-DNA, and HCV-DNA con-
concentration of HAV-DNA, HBV-DNA, and HCV-DNA was
centration. Fig. 3B showed a good linear relationship between
192 nM, 192 nM and 128 nM, respectively. The concentration of
the fluorescence intensity ratio F/F0 (F and F0 refer to the flu-
GO was 30 lg mL 1. The concentration of base mismatched
orescence intensity of the system in the presence and absence of
ssDNA was the same as the target DNA.

Table 1 Comparison of different methods for the determination of hepatitis virus DNA.
Method Material Target Linear Detection Ref.
DNA range limit
Fluorescence polarization SiO2 NP–DNA/Ag NCs HBV 1–200 nM 0.65 nM Chen et al., 2015
Fluorescence detection NH2-UCNPs and Au NP HBV 0–50 nM 0.25 nM Zhu et al., 2015
Fluorescence detection Fluorescent dye HCV 50–400 nM 24.57 nM Zeng et al., 2018
Fluorescence detection Gold nanoparticles-coated HBV — 0.65 nM Fakih et al., 2017
polystyrene beads
Fluorescence detection QDs and magnetic nanospheres HBV 0.5–10 nM 0.148 nM Hu et al., 2013
HCV 0.5–8.0 nM 0.077 nM
Electrochemical detection Graphite electrodes HBV 0.18– 0.18 lM Honorato Castro et al.,
1.8 lM 2014
Molecularly imprinted HAV 0.3–95 nM 3.4 PM Luo, et al., 2016
fluorescence sensor HBV 0.3–90 nM 5.3 PM
Electrochemical detection QDs and Au nanoparticles HBV 0.0005– 0.08 pM Liu et al., 2016b
HCV 0.5 nM 0.34 pM
0.001–1 nM
Fluorescence detection ssDNA-QDs/GO HAV 1.0–192 nM 0.46 nM This work
HBV 8.0–192 nM 1.53 nM
HCV 1.0–128 nM 0.58 nM
ssDNA-QDs/GO multicolor fluorescence system 7

Table 2 Recovery for the detection of hepatitis virus DNA in real samples (Mean ± SD; n = 3).
Hepatitis virus DNA Serum samples (nM) Added amount (nM) Detected (nM) (Mean ± SD) Recovery ratio (%) RSD (%)
HAV-DNA Not detected 16.0 14.75 ± 0.64 92.19 4.03
64.0 69.25 ± 3.02 108.20 4.71
HBV-DNA Not detected 16.0 15.06 ± 0.57 94.13 3.56
64.0 67.28 ± 2.74 105.13 4.29
HCV-DNA Not detected 16.0 15.13 ± 0.62 94.56 3.89
64.0 58.87 ± 3.03 91.98 4.74

those of fluorescence methods (Chen et al., 2015; Zhu et al., applicability of the proposed system for diagnosis of viral hep-
2015; Zeng et al., 2018; Fakih et al., 2017). Compared with sin- atitis by screening viral DNA.
gle or double target detection, the constructed probe could
rapidly detect three kinds of hepatitis viral DNA simultane- 4. Conclusion
ously. Although the sensitivity of our constructed system is
inferior to those of some electrochemical methods and molec- By virtue of the multi-color fluorescence properties of QDs, as well as
ularly imprinted fluorescence sensor (Honorato Castro et al., the unique properties of GO including selective adsorption of ssDNA
2014; Liu et al., 2016b; Luo et al., 2019), modifier preparation and efficient fluorescence quenching properties, ssDNA-QDs/GO mul-
and electrode modification in these strategies are sophisticated ticolor fluorescence system was constructed for synchronous screening
and time consuming. In term of the operation process, the pro- of DNA sequences originating from different types of hepatitis virus.
Taking HAV-DNA, HBV-DNA and HCV-DNA as models, the con-
posed system is convenient and time-efficient.
structed system displayed desirable performance for discriminating
and recognizing three kinds of viral hepatitis DNA. Moreover, syn-
3.4. Specificity chronous detection of HAV-DNA, HBV-DNA and HCV-DNA could
be achieved by measuring the change of fluorescence signals of the sys-
The specific performance of the constructed multicolor fluores- tem under a single wavelength excitation. The proposed strategy can
cence system was investigated by recognition of three hepatitis serve as an alternative method for rapid screening of viral hepatitis
through simple steps. In addition, the present work also provides a cer-
virus DNA in a coexistence system containing their single-
tain reference for the high-throughput detection of multiple disease
base, double-base, and triple-base mismatched ssDNA strands.
markers through multi-color QDs and GO or other nanomaterials.
As displayed in Fig. 4, the fluorescence intensity of three hep-
atitis virus DNA was apparently recovered at 525 nm, 585 nm
Author contributions
and 632 nm, which corresponded to high fluorescence intensity
ratio (F/F0). Compared with hepatitis virus DNA, their single-
base, double-base, and triple-base mismatched ssDNA strands All authors contributed to literature research and manuscript
could not induce significantly recovery of the fluorescence. writing.
This is mainly due to the low hybridization capability of
ssDNA towards its base mismatched complementary Declaration of Competing Interest
sequences. The constructed system can effectively distinguish
and screen three kinds of hepatitis virus DNA from numerous The authors declare that they have no known competing
ssDNA strands. To evaluate the anti-interference of the pro- financial interests or personal relationships that could have
posed multicolor fluorescence system, the effect of some coex- appeared to influence the work reported in this paper.
istent substances (eg. biomolecules and ions) in the biological
fluids on the detection of hepatitis virus DNA was investi- Acknowledgements
gated. As displayed in Table S2, 100 lM Na+, K+, and
Ca2+ can hardly affect the detection of three hepatitis virus This work was supported by the Natural Science Foundation
DNA. Moreover, 200 lM L-Cys, L-Gly, ATP, and BSA also of Shanxi Province of China (NO. 201901D211390) and Nat-
had no obvious effect on the detection results. All of the obser- ural Science Foundation of Shanxi Normal University.
vations above clearly indicated the high selectivity of the pro-
posed method. Appendix A. Supplementary material

3.5. Screening of hepatitis viral DNA in serum sample Supplementary data to this article can be found online at
https://doi.org/10.1016/j.arabjc.2023.104582.
In order to investigate the applicability of the proposed sys-
tem, spiked recovery experiments were performed by adding References
three kinds of hepatitis virus DNA with two concentrations
into the diluted serum sample. The results were presented in Adegoke, O., Forbes, P.B.C., 2016. l-Cysteine-capped core/shell/shell
Table 2. The recoveries for HAV-DNA, HBV-DNA and quantum dot–graphene oxide nanocomposite fluorescence probe
HCV-DNA were 92.19 %-108.20 %, 94.13 %-105.13 %, and for polycyclic aromatic hydrocarbon detection. Talanta 146, 780–
91.18 %-94.56 %. These results demonstrated the potential 788.
8 J. Guo et al.

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