Recent Advances in Biosensors For Detection of COVID-19 and Other Viruses

22 IEEE REVIEWS IN BIOMEDICAL ENGINEERING, VOL.
16, 2023
Recent Advances in Biosensors for Detection

of COVID-19 and Other Viruses
Shobhit K. Patel , Senior Member, IEEE, Jaymit Surve , Graduate Student Member, IEEE,
Juveriya Parmar , Graduate Student Member, IEEE, Kawsar Ahmed , Student Member, IEEE,
Francis M. Bui , Member, IEEE, and Fahad Ahmed Al-Zahrani
(Methodological Review)
Abstract—This century has introduced very deadly, dan- healthcare, wearable electronics, safety, environment, mili-
gerous, and infectious diseases to humankind such as the tary, and agriculture. We strongly believe that these insights
influenza virus, Ebola virus, Zika virus, and the most in- will aid in the study and development of a new generation
fectious SARS-CoV-2 commonly known as COVID-19 and of adaptable virus biosensors for fellow researchers.
have caused epidemics and pandemics across the globe. Index Terms—Biosensor, COVID-19, Ebola, influenza,
For some of these diseases, proper medications, and vacci- pandemic, rapid detection, zika.
nations are missing and the early detection of these viruses
will be critical to saving the patients. And even the vaccines
are available for COVID-19, the new variants of COVID-19 I. INTRODUCTION
such as Delta, and Omicron are spreading at large. The
HE 21st century has introduced very deadly, dangerous,
available virus detection techniques take a long time, are
costly, and complex and some of them generates false neg-
ative or false positive that might cost patients their lives.
T and infectious diseases to humanity such as influenza virus,
Ebola virus, Zika virus, and the most infectious severe acute
The biosensor technique is one of the best qualified to ad- respiratory syndrome coronavirus (SARS-CoV), and SARS-
dress this difficult challenge. In this systematic review, we
have summarized recent advancements in biosensor-based
CoV2. These viruses caused epidemics and pandemics that
detection of these pandemic viruses including COVID-19. humankind has never faced before. But the pandemics that we
Biosensors are emerging as efficient and economical an- are experiencing in the 21st century have their roots in the mid
alytical diagnostic instruments for early-stage illness de- or late 20th century. These viruses have been around since the
tection. They are highly suitable for applications related to 20th century and evolving, and in some scenarios, they have
aggravated [1], [2], [3]. Even though the current epidemics in
Manuscript received 21 March 2022; revised 28 June 2022; accepted South Africa are disastrous, the Ebola virus first appeared in the
23 September 2022. Date of publication 5 October 2022; date of cur-
rent version 6 January 2023. This work was supported in part by the 1970 s. In the same manner, the Zika virus was first identified in
National Science, Technology and Innovation Plan (MAARIFAH), the Uganda in febrile rhesus macaque monkey in the Zika forest and
King Abdul-Aziz City for Science and Technology (KACST), Kingdom later detected in Aedes Africans mosquitoes in the same forest
of Saudi Arabia by under Grant 12-INF2970-10, and in part by the Nat-
ural Sciences and Engineering Research Council of Canada (NSERC). [4]. In 1954 the first 3 human cases were detected in Nigeria
(Corresponding Author: Kawsar Ahmed.) [5] and later it was at large in Brazil from 2015 to 2017 and
Shobhit K. Patel is with the Department of Computer Engineering, was reported in 87 countries by late 2019 across the world.
Marwadi University, Rajkot 360003, India (e-mail: shobhitkumar.patel@
marwadieducation.edu.in). Even the latest COVID-19 pandemic has its first appeared in
Jaymit Surve is with the Department of Electrical Engineer- the 1960 s and later more than 30 variants emerged as Human
ing, Marwadi University, Rajkot 360003, India (e-mail: jaymit.surve@ coronavirus OC-43 (HCoV-OC43), SARS-CoV, HCoV- NL63,
marwadieducation.edu.in).
Juveriya Parmar is with the Department of Mechanical and Materials HCoV-HKU1, Middle East Respiratory Syndrome coronavirus
Engineering, University of Nebraska - Lincoln, Nebraska 68588 USA, (MERS-CoV), and the recent COVID-19 has spread worldwide
and also with the Department of Electronics and Communication En- at large compared to previous coronaviruses [6], [7], [8].
gineering, Marwadi University, Rajkot 360003, India (e-mail: juveriya-
[email protected]). Reverse transcription polymerase chain reaction (RT-PCR),
Kawsar Ahmed is with the Department of Electrical and Computer cell structure, and fast antigen testing are among the major H1N1
Engineering, University of Saskatchewan, Saskatoon, SK S79 5A9, medical testing used in hospitals and healthcare facilities. A
Canada, and also with the Group of Bio-PhotomatiX, Department of In-
formation and Communication Technology, Mawlana Bhashani Science cotton swab is utilized to collect a sample from the nasopharynx,
and Technology University, Santosh, Tangail 1902, Bangladesh (e-mail: which is then used for diagnosis in all three techniques [9].
[email protected];[email protected]). RT-PCR and cell structure are extremely sensitive and precise,
Francis M. Bui is with the Department of Electrical and Computer
Engineering, University of Saskatchewan, Saskatoon, SK S79 5A9, unfortunately, they take much time to validate and require
Canada (e-mail: [email protected]). costly instruments [10]. The quick antigen assay is affordable
Fahad Ahmed Al-Zahrani is with the Department of Computer Engi- and can diagnose a patient in half hour, but it is less robust
neering, Umm Al-Qura University, Mecca 24381, Saudi Arabia (e-mail:
[email protected]). and precise, and it can’t be utilized for validation [11], [12],
Digital Object Identifier 10.1109/RBME.2022.3212038 [13], [14]. Owing to the low amounts of viremia existing at
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PATEL et al.: RECENT ADVANCES IN BIOSENSORS FOR DETECTION OF COVID-19 AND OTHER VIRUSES 23
the beginning of indicators, even screening procedures based [27]. Droplets of varying sizes are the primary means by which
on RT-PCR technology might produce false negative results the COVID-19 infection spreads. In the beginning, the SARS-
within several days of infectivity [15], [16]. The longer an CoV2 virus causes a severe respiratory infection that mimics
individual’s diagnosis is prolonged, the higher percentage of acute respiratory syndrome. Histopathological approaches must
the virus is present in their body fluids, increasing the chance be used to identify COVID-19 in samples with a high risk of
of infection. The PCR test is the standard method for SARS- infection [28]. RSV and HRV cause infants’ respiratory epithe-
CoV-2 identification; nevertheless, it is prohibitively expensive, lium to manufacture inflammatory interleukins (ILs). This study
requires specialized laboratories and expert personnel, and takes compared RSV-negative and RSV-positive patients’ interleukin-
longer [17]. Biosensors can be fabricated by employing var- 8 levels. RSV-positive patients may have higher IL8 levels than
ious techniques that include sputtering, lithography, and the RSV-negative ones; IL8 production tends to be released into the
latest printing techniques, and other than these chemical va- nasopharyngeal region, and the evaluation approach can also
por deposition, electrodeposition, electrochemical exfoliation, alter outcomes [29].
self-assembly, and electrospinning are some of the well-known In this review paper, we have presented an in-depth and
fabrication techniques [18]. Furthermore, each fabrication tech- comprehensive survey of pandemic viruses including the lat-
niques have their advantages and limitations. But among these est COVID-19 virus, H1N1 influenza, Ebola virus, and Zika
techniques lithography is widely used for its accuracy but it virus detection using the rapid detection technique of biosens-
requires expensive systems for nanoscale fabrication and new ing in sections 2 to 5. We have included the recent advances
masks when featuring design changes. And then there is self- in virus detection that includes thorough data related to the
assembly which can provide high resolution with inexpensive- field-effect transistor biosensors, electrochemical biosensors,
ness so we can conclude that self-assembly is an easy fabrication graphene-based biosensors, nanomaterials-enabled biosensors,
technique [19]. label-free biosensors, nanophotonic biosensors, a fiber-optic
As a result, developing rapid, repeatable, commercial, user- biosensor, luminescent biosensors, DNA-based biosensors, and
friendly, precise, and early detection of all these pandemic some hybrid biosensors that provide a rapid, accurate, sensitive
viruses in various specimens is critical. So, it is clear that and low-cost diagnosis of pandemic viruses. The concluding
the rapid, accurate, efficient and low-cost detection of these remarks are presented in section 6. The insights under this review
pandemic viruses will help the medical personnel to start the paper are intended to serve as a foundation for future biosensors’
treatment as early as possible as time is the key factor for saving development for all viral diseases.
the patient’s life. Tremendous efforts are being made around the
world to overcome this constraint by designing suitable, rapid,
and consumer-friendly diagnostic tests that can be used at the
point of necessity [20], [21], [22]. The biosensor technique is II. COVID-19 DETECTION
one of the greatest qualified to address this difficult challenge. In December 2019, a string of pneumonia cases with no known
Biosensors are comprehensive devices that combine explicit etiology was recorded in Wuhan, China’s Hubei province [30].
antibodies, enzymes, or DNA strands with an electrochemical, Later, the 2019 novel coronavirus (2019-nCov) was discovered
optical, or mechanical transducer in such a means that once in a patient’s bronchoalveolar lavage fluid [31]; the International
the targeted analyte interacts with the bioreceptor, the trans- Committee on Taxonomy of Viruses [32] labeled it severe acute
ducer produces a sequence of physicochemical variations that respiratory syndrome coronavirus 2 (SARS-CoV- 2) On March
can be decoded into comprehensible and assessable signals for 12, 2020, the World Health Organization (WHO) declared the
numerical analysis. In this context, biosensors play a major COVID-19 outbreak a pandemic because to the rapid growth
role in the detection of these viruses as it has a significant in human-to-human transmission [33]. Since the first case was
advantage, including a short processing time and excellent se- recorded in Nov 2019, the novel-coronavirus COVID-19 has
lectivity for a variety of biological species. Several biosensor spread to 188 countries and 25 territories all over the globe,
applications are discussed in detail by Patel and co-authors regardless of the combined efforts of WHO and governments to
[23], [24]. control the infection [34]. Since the pandemic started, SARS-
Different immobilization procedures can create biosensors. CoV-2 has also mutated and formed into several variants as it
Biosensors have boosted interest in immobilization techniques. is the nature of the RNA virus. According to WHO, COVID-19
Biosensors used polyphenol oxidase (PPO). Banana Polyphe- had been confirmed in over 537 million instances around the
nol oxidase was purified and immobilized on chitosan-gelatin. world as of June 20, 2022, resulting in 6.32 million deaths
Comparing immobilized and free enzyme characteristics [25]. [35]. We have seen the many deadly variants of COVID-19
As a result of the evolution of receptors for biological com- including Alpha, Gamma, Delta, etc. Even if the vaccines are
pounds over millions of years, scientists are currently designing available but still the danger is not over yet. A new variant of
hybrid devices using nanoscience and biology, known as Nano- COVID-19, named Omicron is found in multiple countries that
biosensors so that they can detect molecules in extremely small are more infectious and spreading rapidly all over the world since
concentrations, inaccessible regions, and even within cells [26]. Nov 2021. All the variants of concern, variants of interest, and
Small RNAs (miRNAs) have been demonstrated to have regu- variants under monitoring are presented in a table form in Table I.
latory effects on a variety of cellular processes and pathways, Mahmud et al. implemented a transcriptomic analysis to identify
including metabolism, viral replication, and cell development COVID-19 [36].
24 IEEE REVIEWS IN BIOMEDICAL ENGINEERING, VOL. 16, 2023
TABLE I Taz and co-authors mainly focused on detecting gene ex-

MUTATED STRAINS OF COVID-19 DESIGNATED AS VARIANTS OF INTEREST
(VOIS) AND VARIANTS OF CONCERN (VOCS) AND VARIANTS UNDER
pression profiles for epithelial cells of human lungs caused
MONITORING (VUMS) [37] by SARS-CoV-2 infections and carried out a genomic analy-
sis to confirm the genomic variance between SARS-CoV and
SARS-CoV-2 [52]. Zamzami fabricated a carbon nanotube
field-effect transistor-based biosensor with high selectivity with
SARS-CoV-2 S1 antigen detection in the 10 mM Ammonium
acetate buffer with a limit of detection of 4.12 fg/mL [53].
A. Electrochemical Microbiosensor Based Detection

Electrochemical-based biosensors are highly utilized given
their ease of access, precision, rapidity, a very low limit of detec-
tion (LOD), and high sensitivity-like features for the detection
of the biomarker regardless of any pretreatment requirement
[54], [55], [56]. Electrochemical biosensors with highly sensi-
tive and selective features were developed by applying the Au,
and Ag-based nanomaterials to update the traditional electrode
design [57], [58], [59], [60]. El-said et al. developed the highly
sensitive and selective Au micropattern-based highly uniform
electrochemical biosensor with increased conductivity for the
detection of COVID-19 [61]. The cyclic voltammetry (CV) and
square wave voltammetry (SWV) schemes based on COVID-19
monitoring are presented that utilize COVID-19 antibodies as
probes. This biosensor detected a wide series of compositions
of COVID-19 protein from 5000 pmol/L to 0.1 nmol/L with
a LOD of 0.276 pmol/L using the square wave voltammetry
method. The real-time usage of this biosensor is validated by
obtaining the sample from a human’s nasal swab, without any
requirement of sample preprocessing.
A dynamic model of the COVID-19 electrochemical biosen-
sor is shown in Fig. 1(a). To obtains the COVID-19 protein, we
Starting from Omicron’s appearance, it had spread world- first integrated the antibody onto a Gold pattern and employed it
wide; increased hospitalizations; demonstrated greater sever- as a particular probe for S protein. The Atomic Force Microscope
ity in young people; attacked defense mechanisms of natural (AFM) technique is also used to investigate the topography of
immunity; was not resistant to available vaccines. To manage this substrate, revealing the production of a uniform gold pattern
COVID-19 effectively and successfully, available options had with a width of about 500 nm and a length of about 1 μm, as
to be efficient in the face of these difficulties [38]. SARS-CoV-2 shown in Fig. 1(b). Raman spectroscopy is being used to confirm
transmits by respiratory droplets, aerosols, airborne, and par- the direct implantation of 50 μM antibody onto the gold pattern.
ticulate matter. Nano-enabled photoelectrochemical oxidation The efficiency of the proposed SWV-based sensor was also
(PECO) assisted air purification is a good technological alter- investigated. Fig. 1(c) depicted the square wave voltammetry
native [39]. Intelligent sensor systems combined with IoTs and voltammograms of an antibody/Gold pattern electrode after
AI techniques with 2D nanomaterials have transformed sensor interrelating with various portions of COVID-19 protein ranging
applications in wearable electronics, healthcare, environment, from 5000 fmol/L to 0.1 μmol/L. The value of current is strength-
safety, defense, and agriculture [40], [41], [42], [43], [44], [45], ened as the protein concentration rises, according to the research.
[46], [47]. Due to new SARS-CoV-2 variations, the severity of Fig. 1(d) depicted the link between COVID-19 protein portions
COVID-19 has worsened, causing health authorities to reeval- and the associated current peak at about 450 mV, which exhibited
uate pandemic management techniques. This is crucial due to linear plots with a slope of 3.22 and R2 of 0.988 within this
widespread infection and the global healthcare system’s flaws concentration range. The developed sensor’s LOD was found
[48], [49]. Low-level detection of a selected disease biomarker to be 276 fmol L−1 using the SWV approach. The real-time
(pM level) is valuable for monitoring disease development. detection was also followed by first confirming COVID-19 using
Bioinformatics and multi-aspect-oriented analytics are needed PCR validating the presence of the RNA gene of COVID-19
to examine treatment effectiveness, optimize therapy, and cor- that obtained the oxidation peak at 300 mV and a cathodic
relate biomarker levels with illness pathogenesis [50]. Ahmadi- peak at 100 mV. To conclude proposed biosensor exhibited high
vand et al. focus on making a tiny plasmonic immunosensor sensitivity toward COVID-19 protein and is capable to detect
based on toroidal electrodynamics that can sustain ultranarrow COVID-19 in human specimens without any complex specimen
plasmonic modes at THz frequencies [51]. processes.
by the fabrication of semiconductors are some of the technol-

ogy’s main advantages. These features make this technology
appropriate for multiplexed investigation, where the analytical
value enhances as the panel size grows. In addition, unlike
many common approaches, which use endpoint-based detection
methodologies, real-time surveillance speeds up test progress
and allows for direct inspection of biological intellection [65],
[66].
A BiMW biosensor’s schematic diagram is available in
Fig. 1(e). Conventional silicon microelectronics technology is
used to manufacture the BiMW sensor chip. Waveguides of
micro/nano dimensions are used in the BiMW technology’s de-
sign. The materials were chosen for their strong refractive index
contrast, which is required for a highly sensitive waveguide-
based transducer. The BiMW sensor surface is customized with
particular receptors targeting exterior antigens of the virus, such
as the Spike (S) protein of COVID-19 as shown in Fig. 1(f).
The virus traces are acquired by the receptors on the sensor
surface once the card is inserted into the biosensor, resulting in
an interferometric signal that is logged in real-time. Because the
sensor response is proportionate to the composition of viruses
in the specimen, the viral load in the patient may be accurately
quantified. The bioreceptors must be chosen carefully since they
provide both the required affinity and specificity to the assay.
While antigen-based tests can be used to screen for and identify
COVID-19 contagion, the comparatively minor changes in viral
proteins across various virus categories may cause biorecep-
tor cross-reactivity challenges. The assay is basic, requiring
just prior RNA abstraction and disintegration from the virus,
which can be implemented quickly after specimen assortment.
After the specimen was injected into the device, complementary
Fig. 1. Various COVID-19 detecting biosensors (a) Illustrative proto- single-stranded DNA (ssDNA) capture the specific target gene
type of COVID-19 electrochemical micro-biosensor including immobi- fragments, establishing a duplex chain and resulting in real-time
lization of antibody and obtaining COVID-19 protein (b) AFM topography
of the gold assembly (c) SWV voltammograms for different compositions signals proportionate to the number of viruses as shown in
of COVID-19 protein from 5000 fmol L-1 0.1 µmol L-1 after interact- Fig. 1(g).
ing antiCOVID-19 electrode (d) the relation among the composition The BiMW biosensor’s high levels of sensitivity indicate that
of COVID-19 protein and oxidation current peak (Reprinted from [61],
copyright Springer Nature) (e) A POC biosensor based on biomedical the unique POC diagnostic equipment will operate efficiently,
waveguide (BiMW) interferometric technology (f) Detection of intact with LOD in the region of 102–103 viruses per milliliter for
SARS-CoV-2 using BiMW with CONVAT technique (g) Viral genomic straight and fast SARS-CoV-2 virus detection. Given that most
analysis using BiMW with CONVAT technique (Reprinted from [62],
copyright IOPSCIENCE). COVID-positive patients have viral counts in the span of 105–
107 viruses per milliliter, this result is a decent indication that the
CONVAT technique may be able to detect even asymptomatic
cases with small viral levels. In terms of the genomic assay, the
B. Nanophotonic Biosensor Based Detection capacity to identify minute amounts of RNA (aM-fM levels) in
A point of care (POC) nanophotonic biosensor for the straight, a straightforward manner in only 30 minutes will considerably
efficient, and precise detection of COVID-19 from both human improve the accuracy of COVID-19 diagnosis in large inhabitant
specimens and animal sources is being developed as part of the testing. The device might be used for routine diagnostic analysis
European CONVAT project [62]. The technique can be used of a wide range of diseases and medical conditions, and it could
in isolated surroundings to enhance initial analysis and patient be employed in hospitals, laboratories, and primary care centers.
management, besides monitoring the coronavirus environment The chips can be mass produced using traditional fabrication
to avoid future epidemics. CONVAT presents a novel biosen- techniques, resulting in an affordable system that is projected to
sor based on interferometric technology, which has previously be under 15 € per assay.
been validated for highly sensitive detection of biomarkers and
pathogens [63], [64]. The main aim of CONVAT project is to
advance a POC biosensor prototype for precise and efficient C. Electrochemical Biosensor Based Detection
detection of the SARS-CoV-2 in a short time even in isolated COVID-19 diagnostic tests can be divided into two cate-
surroundings. The low footprint and great scalability enabled gories: serological and viral nucleic acid assays. The first testing
Fig. 2. Electrochemical biosensor-based nucleic acid testing (a) Brief workflow of the biosensor with N genes and S genes RCA, SEM images
(b) Silica core, silica-methylene blue (SiMB), and silica-acridine orange (SiAO) with various diameter size represented as mean value with standard
deviation (c) Comparison of step-wise and one-step sandwich hybridization process with N gene as the target, No significant difference is
observed in these two strategies with sensitivity test for N and S genes positive correlation for a current response, increment in differential pulse
voltammetry as N and S gene concentration increases (d) Numerous sequence position of N genes and S genes target orders with mismatch and
non-complementary target orders. Dark areas and underlined bases show the non-complementary arrangements to the target gene and mismatch
bases, respectively (e) The identification of N genes and S genes in 55 cDNA specimens equated with the Cq result from qRT-PCR (N genes (Blue
dots), S genes (Orange dots)) (Reprinted from [69], copyright Springer Nature).
determines the existence of antibodies generated by an individ- was applied for the enhanced test because it was comparatively
ual as a result of virus infection or the presence of antigenic viral easy, rapid, and required fewer preprocessing steps. The rise in
proteins in infected patients. Rolling circle amplification (RCA), electrochemical signals was certainly linked with the rise in gene
an isothermal amplification method, has also been frequently copy amount for the S genes and N genes as well (Fig. 2(c)).
utilized for nucleic acid testing [67]. With little reagents, RCA The respiratory infections COVID-19 and influenza are both
can yield amplicons 109-fold in 90 minutes [68]. Chaibun et al. infectious. COVID-19 and influenza have nearly identical symp-
reported an electrochemical biosensor for the rapid detection toms, despite the fact that they are caused by separate viruses.
of the S gene and N gene of SARS-CoV-2 from the clinical It might be difficult to tell the difference only based on med-
specimen as shown in Fig. 2(a) [69]. In less than 2 hours, the ical symptoms. To avoid misclassification, an analytic assay
test could identify as few as 1 copy L-1 of viral N or S genes. that could identify the influenza virus from SARS-CoV-2 is
Clinical specimens were utilized to assess the test’s efficiency, essential. 1 pM of non-complementary target orders of Influenza
which is shown to be consistent with the results of qRT-PCR. A and Influenza B viruses was used in the specificity testing,
Through surface-reactive functional groups, two redox dyes, with the sequence alignment displayed in Fig. 2(d). There was
MB and AO, were coated onto silicon nanoparticles. SEM im- additionally 1 pM of linear targets with two bases mismatch
ages of SiMB, and SiAO are shown in Fig. 2(b). In comparison to added which is also presented in Fig. 2(d). A positive result
the silica core, the size of the silicon nanoparticles after coating was defined as a current signal that was equivalent to or more
with the redox dye increased. The current signal for the two tech- than +3 standard deviations above the average mean of the
niques did not differ significantly as the p-value is less than 0.05 background signal. In RCA, RNA and cDNA specimens pro-
(Fig. 2(c)). As a result, the single-step hybridization approach duced from clinical specimens were employed as the template,
TABLE II
COMPARISON TABLE OF COVID-19 DETECTING SENSORS WITH
PREVIOUSLY PUBLISHED WORK
lucKey and lucCage are combined, luciferase activity is recon-

Fig. 3. (a) Illustrative mechanism of the proposed sensor (b) The ther- stituted. In the absence of a target, the binding energy of Luckey
modynamics behind the activation of sensor (c) Specificity of the pro- to lucCage is not enough to surpass the free energy charge
posed sensor for various cognate targets and targets for other biosen- of lucCage opening, for the tuning of thermodynamics of the
sors (d) Incorporating of de novo SARS-CoV-2 RBD binder, Increment in
luminescence due to the addition of trimeric spike protein, and detection system, as shown in Fig. 3(b). The author’s incorporated sensors
over a range of analyte compositions in buffer (Reprinted from [71], that can directly detect COVID-19 viral particles, as shown in
copyright Springer Nature). Fig. 3(d). As available in Fig. 3(c), each sensor responds quickly.
Table II presents the comparison study of available COVID-19
detecting biosensors based on their limit of detection (LOD) and
with electrochemical detection and one-step hybridization. As other important details.
shown in Fig. 2(e), including 30 cDNA and 11 RNA totaling 41
specimens prepared from COVID-19 +Ve specimen produced
+Ve results, whereas the including 25 cDNA, 40 RNA totaling III. INFLUENZA DETECTION
65 specimens prepared from COVID-19 -Ve specimen produced The influenza virus, often called flu, is an extremely severe
-Ve results. This matched the results of the qRT-PCR. The limit respiratory illness, including the 1918 Spanish influenza H1N1,
of detection (LoD) of the proposed assay demonstrated that it 1957 H2N2, 1968 H3N3, and the 2009 H1N1 influenza has
is incredibly sensitive compared to already accessible assay kits impacted millions of individuals around the globe, and innu-
and prevailing biosensors. merable financial and social costs [89], [90], [91]. Conferring
to the WHO, 2000 million individuals are diseased with flu-like
pathogens comprising Influenza, among these $250 k - $500 k
D. Protein-Based Biosensor for Detection worldwide annual deaths are reported [92]. After that new
Biosensors for cellular and clinical applications have been de- animal-originated influenza viruses such as H5N1, H7N9, and
veloped using naturally occurring protein switches [70]. Authors H9N2 have infected humans on rare occasions but had not been
anticipated that these properties may be achieved by the presence transmitted at large [93], [94]. However, extended mutation of
of a peptide actuator controls They implemented a model that these viruses could steer to full adaption to humans and high and
consists of two protein elements: a ‘lucCage,’ which consists of long-term transmission capabilities. Since its first apparition in
a cage domain and a latch domain containing a target-binding 2013, the influenza H7N9 virus has spread the most, with 1,567
motif and a split luciferase fragment; and a ‘lucKey,’ which laboratory +Ve human infectivity [95]. Since the establishment
consists of a key peptide that ties to the open state of lucCage of the benchmark in virological diagnostics – viral separation
as shown in Fig. 3(a). As illustrated in Fig. 3(a) right side, when from tissue structures or embryonated chicken eggs – multiple
approaches for rapid (< 2 h) detection of the influenza virus have The multifunctional DNA 4WJ has comprised of four ssDNA
been established. The majority of modern approaches rely on molecules that each have a significant characteristic. In the head
PCR (conventional PCR, RT-PCR, etc.) [96], [97], [98], [99] or and tail sections of this 4WJ, the influenza aptamer and amine
immunoassay technologies [100]. PCR technologies have more group were inserted. The DNA aptamer attaches to a specific
sensitivity and specificity than other molecular procedures, and target molecule. Easy adaptation to the needed form, minimal
they take less time [101]. rate of manufacturing via chemical combination, great stability,
and minimal cost production are only a few of the benefits. An
A. Boron-Doped Diamond-Based Biosensor for addition of carboxyl-MoS2 enhanced the sensor’s sensitivity.
Detection Field-emission SEM was utilized to confirm the synthesis of
carboxyl-MoS2, and AFM was used to confirm the immobi-
Nidzworski and co-authors presented a biosensor for the early
lization process of each stage. Furthermore, the detection was
detection of the influenza virus [102]. The findings allow for
found to be possible at concentrations ranging from 0.1 μM
high-precision, high-throughput influenza virus detection of the
to 0.01 nM. The LOD of 0.01 nM was achieved. It was also
virus. The reason behind choosing the M1 (matrix) protein as a
established that this sensor exclusively reacted to H1N1 by
detection target is that it is the sole viral element required for
conducting the selectivity test. Furthermore, studies revealed
the generation of VLPs (virus-like particles) and is found in very
that the proposed bioprobe can be employed in real-world
influenza virus serotypes.
scenarios.
For the precise detection of the influenza virus, a label-free,
highly sensitive biosensor was developed and evaluated. High
sensing capability and great stability are features of the improved C. Label-Free Biosensor for Detection
BDD electrodes. Furthermore, the findings show that the pro-
Many researchers have concentrated on analyzing the impacts
posed method has a number of advantages over existing meth-
of linking architectures of sialic acid clusters in host receptor
ods, including a short detection and incubation time (less than
compound glycans, which differ from class to class because
5 minutes), extremely high sensitivity (LOD 1 fg/ml), stability,
HA receptor explicity is a major aspect in the progression of
and high repeatability in influenza virus detection, and the ability
contagion, spread, and adaptability of influenza viruses [105].
to perform consistently. In comparison to currently utilized an-
The N-acetylneuraminic acid, 2,6, galactose (α2,6 associated
alytical procedures, this technique has the shortest investigation
sialic acid moieties), is presented in Fig. 4(a), which are pre-
time and the lowest LOD. The difference in charge transference
dominantly located on the epithelial units of the human upper
resistance was even small than 10% and was consistent across all
ventilatory tract, exhibit a binding preference for human adapted
verified electrodes. Given the extensive application of this strat-
influenza HA proteins [106]. Animal influenza HA proteins, on
egy, the authors are confident that the current findings will result
the other hand, target the NeuAco2,3 Gal (2,3 linked sialic acid
in significant improvements in medical analytical approaches in
moieties), which are prevalent in epithelial units of the entrails
the near future.
as well as the entire ventilatory system of animals and birds
[107]. Numerous researchers have shown that the alteration in
B. DNA 4WJ Based Electrochemical Biosensor for
the linking architectures of 2,6 vs 2,3 sialic acids impacts the
Detection
capacity of influenza viruses to affect various classes, despite
Park et al.presented a biosensor comprised of 4-way junc- its apparent insignificance [108]. Although PCR is the prime
tion (4WJ) and carboxyl-MoS2 hybrid material to accurately standard for detecting the H1N1 virus, it cannot identify the
detect H1N1 [103]. The hemagglutinin aptamer was placed ventilatory-binding explicity of remote influenza (H1N1) virus
on the recognition part (head group), four silver ions were a priori. As a result, sensors for quickly determining HA ven-
placed on each of the two arms (signal amplification part), and tilatory explicity, particularly the distinction of influenza HA
an amine group was placed on the tail group (anchor). This proteins binding 2,6 and 2,3 associated sialic acids, are crucial
manufactured multifunctional DNA four-way junction can bind for evaluating the adaptative capabilities of newly developing
to hemagglutinin specifically and selectively. Furthermore, the animal influenza viruses.
carboxyl-MoS2 improves the sensitivity of this biosensor. An Label-free sensor systems capable of profiling influenza HA
electrochemical study revealed the existence of H1N1 when it glycan binding explicity in a microarray style are therefore ex-
was inserted into the immobilized electrode. It was also able to tremely required. To meet this prerequisite, Zhang et al. used the
establish whether this sensor responds explicitly and selectively Arrayed Imaging Reflectometry (AIR) sensor platform to pro-
to the influenza virus (H1N1) through selectivity testing. It is also duce and evaluate microarrays of influenza ventilatory-binding
confirmed that the biosensor responded linearly to influenza and analogues [104]. AIR is a multiplex detection system that does
that influenza could be sensed at concentrations ranging from not require the use of labels. The platform makes use of a
0.1 μM to 0.01 nM. Finally, laboratory experiments using human single-camera interferometric imaging setup. A charge-coupled
serum diluted hemagglutinin exhibited an analogous inclination device (CCD) camera captures the array’s reflected image.
to those using water diluted hemagglutinin. The multi-functional Fig. 4(b) depicts the microarray chip processing and polymer-
DNA four way junction and carboxyl-MoS2 hybrid substance based glycan-antigen identification method. The microarray lay-
can be used to make an electrochemical influenza virus (H1N1) out is shown in Fig. 4(c). As hypothesized, influenza H1N1pdm
detecting biosensor, according to this study. coupled to the α2,6 linked sialic acid moieties probe sites with
TABLE III
COMPARISON TABLE OF INFLUENZA VIRUS DETECTING SENSORS WITH
America has caused thousands of deaths that resulted in world-

wide chaos [120]. The Ebola virus is a type A select agent
filovirus that was initially discovered in 1976 in Zaire and named
after the Ebola River [121]. Person-to-person transmission of
Ebola is reflected by direct contact with infected patients, bod-
ily fluids, or compromised clothing. Enhanced disease control
actions and primary medication at specialist institutions might be
possible if this highly contagious virus could be detected early.
The present fatality rate from epidemic Ebola varies from 40%
to 90%, and early, point-of-care diagnoses could substantially
Fig. 4. Label-free, Multiplex microarray biosensor for influenza detec- reduce this number [122], [123]. According to mathematical
tion (a) Human and avian cell surface sialyloligosaccharide structures calculations, the total direct costs of this outbreak in the three
(b) Preparation and incubation of AIR microarray chips for virus-glycan
binding detection (c) Glycan-based receptor analogue microarray layout most affected nations vary from 82 million to 356 million;
design (d) AIR glycosyl microarray responses to H1N1pdm and H13N8 early identification and medication will also minimize overall
viruses at various viral doses (e) The glycan microarray’s quantitative healthcare expenditures [124].
response data to the H1N1pdm virus (left) and the H13N8 virus at right.
(f) Analysis protocol for AIR images (Reprinted from [104], copyright Various ELISA testing, RT-PCR testing, virus separation,
ACS Publications). electron microscopy, and serologic testing for IgM or IgG anti-
bodies are being employed to identify Ebola. Quantitative viral
load testing is also prognostic, with patients with viral loads of
remarkable selectivity. Similarly, the H13N8 virus bonded only more than 10 million copies/mL having a substantially higher
to the NeuAc2,3 Gal linked glycan probe sites, demonstrating case fatality rate [125]. Due to the very small amounts of viremia
that it is species specific. For human and bird influenza viruses, that exist at the outset of indications, even analytic procedures
quantified response data approves explicit binding of the virus to based on the RT-PCR method might provide false negative
individual glycan probe as shown in Fig. 4(e). As illustrated in outcomes in the first few days of infectivity [126]. The longer a
Fig. 4(d), the AIR microarray pictures were collected, recorded, patient’s diagnosis is delayed, the more virus is existing in their
and examined for quantitative responses. The response values body liquids, increasing the chance of infection. EVD can also
for each array location were determined using an empirically have a long incubation period, up to three weeks, necessitating
resultant response model to convert the reflection intensity unit the use of diagnostic tests. Several diagnostic approaches have
to thickness. The response model depicts the connection among been investigated for quick diagnosis of EVD.
reflected intensities measured by ellipsometry and their corre-
sponding thicknesses for various exposures. Fig. 4(f) shows an
A. Surface Acoustic Wave (SAW) Biosensor for
example of this technique employing a chip containing the α2,6
Detection
glycan analogue and treated with SNA lectin. Table III presents
the comparison study of available Influenza virus detecting Rapid Ebola virus diagnosis at the point of care could allow
biosensors based on their limit of detection (LOD) and range for early containment and the averting of future outbreaks and
of functionality. pandemics [127]. Although there are POC nucleotide intensi-
fication tests, they are inadequate by the necessity for several
reagents, preservation, and expert persons. Baca and co-authors
IV. EBOLA DETECTION
presented initial outcomes of a SAW biosensor that has the abil-
The Ebola virus disease (EVD) epidemic of 2014-15 in West ity to detect the EVD within 5–10 minutes [128]. SAW sensors
African countries and infrequent cases in Europe and North can identify EVD antigens fast at the POC, with no additional
chemicals, sample processing, or trained people required. The

authors utilized a deactivated virus to proceed with this work
under non-BSL-4 settings because the fully intact Ebola virus is
very infectious.
Prior to virus inactivation, EVD antigens were found in
phosphate-buffered saline (PBS) solutions at concentrations
ranging from 1 × 104 PFU/mL to 3 × 106 PFU/mL. On the first
day of illness symptoms, Ebola patients had an average viremia
level of 30 RNA kcopies/mL [129]. EVD detection occasioned
in a composition-dependent rise with phase shit values roughly
spaning from 16 kPFU mL−1 to 6.5 MPFU mL−1 . By employing
linear regression and the other factors, an Ebola virus limit of
detection of 19 kPFU/mL was achieved. These findings show
that the prototype SAW biosensor identifies EVD antigens in a
specified buffer fast, with LOD under the mean viremia level at
the beginning of medical indications.
B. Electrochemical DNA Biosensor for Detection

Many clinical and environmental investigations have focused
on ssDNA identification, such as cancer diagnosis [130], bac-
terium detection [131], SPR [132], and spectroscopic methods
approaches [133]. Electrochemical DNA biosensors stand out
because of their distinct advantages, which include affordable,
quick reaction, user-friendly, correct selectivity, and high sensi-
tivity.
Ilkhani et al. developed a new electrochemical DNA biosensor
for detecting the Ebola virus [134]. The electrochemical biosen-
sor was created by applying S-Au boundaries. The hybridization
process was detected using EIS and DPV. Scrutinized using the
electrochemical impedance spectroscopy approach and used as
a signal for label-free DNA hybridization exposure. The results
showed that the EVD DNA can be identified with great re- Fig. 5. (a) Illustrative diagram of reduced graphene oxide-based FET
biosensor (b) SEM image of reduced Graphene Oxide sheet (c) AFM
peatability, sensitivity, and selectivity using an electrochemical used to determine the depth of the reduced Graphene Oxide sheet,
biosensor. As an analytical technique, this system can be used to which was found to be less than 2 nm (d) EGP inserted in 10−3 ×
detect Ebola virus DNA in genuine samples. This approach can human serum (e) EGP inserted in 10−3 × human plasma (f) Detection
of MARV GP in PBS (g) Sensitivity as a function of protein composition
also be used to analyze bacterial pollution in the environment, is investigated. (Reprinted from [135], copyright Springer Nature).
such as in water or food sources.
C. Field-Effect Transistor (FET) Biosensor for Detection Anti-Ebola probes have been mounted on the channel, and they
For real-time detection of the EVD antigen, Chen et al. preferentially catch the antigen. The authors discovered that such
developed a rGO FET technique [135]. This technology takes a FET biosensor exhibits great fundamental characteristics for
advantage of graphene’s appealing semiconductor properties the Zaire strain’s Ebola GP, with a detection limit of 1 ng/ml.
to produce a very sensitive and explicit identification of EVD The Ebola antibody is mounted to identify the particular
glycoprotein in real time. These findings show that an enhanced antigen in Fig. 5(a), which depicts an illustrative structure of
FET biosensor for EVD diagnostics may be successfully fabri- the reduced Graphene Oxide-based biosensor mechanism. The
cated. The FET is a potential approach for detecting a variety design of the produced biosensor device was investigated using
of analytes quickly and accurately. Its applicability has been SEM as shown in Fig. 5(b). Sputter coating was used to deposit
established in the detection of target analytes in gases [136] and GNPs homogeneously on the mechanism for antibody conju-
water [137], for example. Rapid reaction, minimal cost, and ease gation. The gold NPs weren’t visible prior to sputter coating,
of use are all advantages of FET sensors. By affixing explicit implying that these glowing “dots” are GNPs created by the
probes on the conducting channel, FET biosensors can attain sputter coater. The depth of the reduced Graphene Oxide sheet
improved selectivity and sensitivity, defined as a fundamental is calculated using AFM to better characterize the FET device.
feature for FET sensor accomplishment. When compared to The reduced Graphene Oxide sheet has a depth of roughly 2 nm,
graphene prepared by the CVD approach, rGO FET devices may demonstrating a few-layer structure, while the monolayer depth
be simply made by thermal annealing graphene oxide sheets, is 340 μm, as seen in Fig. 5(c). The authors utilized 10−3 ×
with the added benefits of affordable and empirical pliability. human serum per plasma acquired from the Blood Center of
TABLE IV
COMPARISON TABLE OF EBOLA VIRUS DETECTING SENSORS WITH
the sensitivity of EGP, avidin, SUDV GP, and MARV GP as a

function of composition with error bars. Table IV presents the
comparison study of available Ebola virus detecting biosensors
based on their limit of detection (LOD), techniques used for
detection, processing time, and other important details.
V. ZIKA DETECTION
The first substantial outbreak of Zika fever occurred on the
Fig. 6. Gold nanoparticles based electrochemical biosensor based
Zika virus detection (a) Illustration of the concept to amplify the Federated States of Micronesia’s Western Pacific Island of Yap
nanobioconjugate-based signal (b) Assessment of molar ration using in 2007 [143]. Multiple incidences of ZIKV have been recorded
anti-Dig-HRP as a reporter (c) For various gold and NaCl composi- since then all across the world. In French Polynesia, two major
tion, chronoamperometry curves for the gold electrodeposition (d) SEM
images of gold resultant structures at various enlargement (e) DPV crises impacting over 30,000 residents occurred in 2013 and
plots of nanobioconjugates accumulated onto SPGEs and (f) Resultant 2014 [144], [145]. Smaller epidemics were also reported in New
calibration curve (g) DPV plots of three positive ra serum specimens (h) Caledonia, Easter Island, and the Cook Islands, as well as the
Matrix effect and specificity (Reprinted from [150], copyright Springer
Nature). Solomon Islands, Samoa, and Vanuatu. Later, it was reported
in continental South America in Brazil in May 2015 and WHO
declared it as a public health emergency in February 2016 as
Wisconsin to suspend Ebola glycoprotein as a technique to repli- 1.3 million people were infected in Brazil itself. Furthermore,
cate blood specimen from EVD +Ve persons to examine sensor more than 20 countries in the Americas have reported the au-
performance under a more difficult yet real-time environment. tochthonous Zika virus transmission, comprising Puerto Rico
The sensor’s dynamic response to EGP in 10−3 × serum is and US Virgin Islands during 2016 [146].
shown in Fig. 5(d). As the concentration of EGP increased, The US Food and Drug Administration has approved 14
the Id decreased as well. The biosensor was then refilled with rRT-PCR screening assays and 5 anti-ZIKV IgM serological
0.01 × serum, but no substantial variation occurred, demon- tests. For a point of care testing (POCT), only one rRT-PCR and
strating that only explicit binding can cause a current variation. four serological screening assays are utilized, with a sensitivity
Fig. 5(e) shows the dynamic response to EGP in 0.01 × plasma, and specificity of more than 90%. These assessments, however,
with respect to a significantly noisier signal. The serum/plasma take roughly 1–3 hours to complete [147]. Cross-reaction of
reaction to EGP was similar to that in PBS, but with a decreased ZIKV antibodies particularly with DENV, is possible, thereby
sensitivity (about 1.7% for 1 μg L−1 Ebola glycoprotein in increasing false positives in several immunoassays [148]. Fur-
10−3 × serum), demonstrating the complex dispersion medium thermore, serological and molecular assays necessitate costly
affects the response. Fig. 5(f) depicts the dynamic responses instruments and skills to operate. The majority of ZIKV in-
to non-specific GP. The sensitivity of 1 ng/ml of MARV GP fections occur in developing nations, which lack adequate lab-
is just about 0.18 percent. As a result, the FET biosensor has oratory facilities, skilled personnel, and financial resources.
a low reaction to non-explicit glycan proteins and significantly As a result, fresh approaches are needed to develop a ZIKV-
lower sensitivity than the EGP, implying that cross reactivity is specific POC scheme that can distinguish between ZIKV and
low and the sensor has good selectivity for the target protein. DENV infections in epidemic areas that are efficient, low-cost,
In Fig. 5(g), five independent replicates are used to compare and quick.
A. Peptide Aptamer Pair-Linked Based Biosensor for TABLE V

COMPARISON TABLE OF ZIKA VIRUS DETECTING SENSORS WITH
Detection PREVIOUSLY PUBLISHED WORK
To identify ZIKV, Nguyen et al. created a new biosensor
[149]. A peptide aptamers pair (PAP) were used to create an
antibody-free lateral FICT. The LOD for the B2.33-P6.1 PAP
for ZIKV in the quick diagnostic strip was 20000 tissue culture
infective dose TCID50 /mL. Significantly, FICT was able to
distinguish ZIKV from DENV. Human sera and urine were
used to confirm the stability and performance of FICT, and the
LOD value was found to be equivalent. In silico modeling was
employed to design a novel PAP FICT assay for identifying
ZIKV, according to the study.
The TL/CL value was used to calculate the LOD of FICT. At
8 × 104 TCID50 /mL, P29.1 and Z10.8 , strongly distinguished
ZIKV. ZIKV was detected at a higher titer, 320000 TCID50 /mL,
by the -Ve Z10.2 peptide in combination with B2.33, which was
substantially dissimilar from the +Ve peptides. With satisfactory
linear regression, the ZIKV titer related to the limit of detection
was 40000 TCID50 /mL, while the limit of detection was 20000
TCID50 mL−1 . P6.1 had a considerably greater signal for ZIKV
identification with p value less than 0.001, demonstrating that
the novel P6.1 peptide detects ZIKV better than the earlier Z10.8
peptide, which is a constant with predictions. Peptide aptamers
have been successfully produced by Authors as a useful diag-
nostic material for distinguishing between ZIKV and DENV.
This innovative PAP FICT test could well be conducted in 20
minutes and used as a POC scheme to detect ZIKV in human
serum and urine.
B. Gold Nanoparticles Based Electrochemical Biosensor

for Detection
A durable nanobioconjugate relied upon gold nanoparticles
(GNPs) connected to ssDNA has been developed for high-
sensitivity magnification of the electrochemical signal of a ZIKV
[150]. The genosensor is designed with Ru3+ as an electrochem-
ical reporter on a screen-printed gold electrode (SPGE)/screen-
printed carbon electrode (SPCE) designed with layered gold
nanostructures (SPCE/G). The genosensor performance was aM. Serum samples from diseased individuals showed variable
straight from 0.01 to 0.6 aM and from 0.5 aM to 0.01 fM of current density responses. In comparison to the -Ve control
the targets, as measured by DPV, with a sensitivity of 2700 (NC1) with no target, serum specimens from diseased patients
and 2900 mA cm2 M−1 and a LOD of 0.2 and 33 fM at the demonstrated variable current density outcomes, which might
SPGE and SPCE/G, accordingly. Nanobioconjugates are hybrid be connected to the RNA loading in every specimen (Fig. 6(g)).
nanomaterials formed when nanomaterials and biomolecules As depicted in Fig. 6(h), the electrochemical sensor was
are combined [151]. Hundreds of active biomolecules and tags also evaluated in urine and saliva samples doped with 1000
have been immobilized on top of nanomaterials to boost signal fM synthetic explicit DNA. The research indicated that the
strengthening responsiveness. The conjugation of GNPs with signal strength in the urine sample was greater than that in
ssDNA provides a simple way to synthesize nanobioconjugates the saliva specimen, even when the target concentration was
with exceptional capabilities for signal amplification, improved the same. Table V presents the comparison study of available
sensitivity, and lower LOD in bioassays. A solution in the Zika virus detecting biosensors based on their limit of detection
existence of sodium chloride is shown in Fig. 6(d) and the inset (LOD), techniques used for detection, processing time, and other
at various enlargements. The DPV plots for highly sensitive important details.
identification of the Zika synthetic genetic substance formed at This is the first GNP-ssDNA nanobioconjugate to be em-
the exterior of SPGEs are shown in Fig. 6(e). With a sensitivity of ployed in the ultrasensitive recognition of ZIKV genetic sub-
2900 nA cm2 M−1 and a LOD of 0.2 fM as illustrated in Fig. 6(f), stance without the use of specimen abstraction or PCR. The
they exhibit a linear dependency of the electrochemical response gene-based assay was developed with a Ru3+ complex as an
with the increasing target composition in the range of 0.01 to 0.6 electrochemical reporter on either a SPGE or a SPCE modified
with Au layered nanostructures (SPCE/G) as shown in Fig. 6(a). REFERENCES

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Recent Advances in Biosensors For Detection of COVID-19 and Other Viruses

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22 IEEE REVIEWS IN BIOMEDICAL ENGINEERING, VOL.

Recent Advances in Biosensors for Detection

TABLE I Taz and co-authors mainly focused on detecting gene ex-

A. Electrochemical Microbiosensor Based Detection

by the fabrication of semiconductors are some of the technol-

lucKey and lucCage are combined, luciferase activity is recon-

America has caused thousands of deaths that resulted in world-

chemicals, sample processing, or trained people required. The

B. Electrochemical DNA Biosensor for Detection

the sensitivity of EGP, avidin, SUDV GP, and MARV GP as a

A. Peptide Aptamer Pair-Linked Based Biosensor for TABLE V

B. Gold Nanoparticles Based Electrochemical Biosensor

with Au layered nanostructures (SPCE/G) as shown in Fig. 6(a). REFERENCES

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