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Файл:Ординатура / Хирургия / Библиотека им академика М.И. Перельмана / Книга_5361_Библиотеки_им_академика_М_И_Перельмана.pdf
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A. Chakraborty et al.
label- free mechanics used, the ease of miniaturization, low cost and easy processing
of electrical biosensor make them the point-of-care in disease diagnostics [9].
There are different transducer mechanisms used in biosensors like electrochemical, optical, thermal and calorimetric [10]. While chemiluminescent, absorbance,
and uorescence are basic mechanisms behind optical transducer signal output, the
mechanism of the electrochemical transducer biosensor is generally conductometric
[11, 12]. Based on the nature of the transducer used, the signal is received from the
bioreceptor and further conveyed to the processor. In between this, the signal is
amplied while the output amplitude is proportional to the analyte’s concentration
[13]. On binding of the analyte to the bioreceptor, a change in physiochemical signal is captured and converted to electrical signal by transducer. Analysis of the
changes in different parameters of the signal like conductance, intensity, and potential reveals the presence or absence of bioagent [7]. Apart from biotransducer, there
are other vital components of a biosensor, microprocessor, monitor and most important, the bioreceptor, which is specic for biorecognition, to detect the analyte
[14, 15].
In spite of all these advantages, there are few hurdles that prevent the wide-scale
use of afnity biosensor in disease diagnostics. First is multiplexing, i.e., the ability
to detect a wide range of analytes in a single sample. The second obstacle is the high
detection limit. These problems can be overcome by the implementation of
nanoscale biosensor as the potential candidate, with increased sensitivity and lower
detection limit [16]. The use of nanotechnology allows manipulation at the atomic
level where the dimension remains in the range of 1–100 nm scale [17]. Their
extremely small size improves performance in electrochemical and enzymatic biosensors by increasing the electron transfer rates as well as by shortening enzyme-toelectrode distances [18].
Nanosensors have the special advantage of displaying signicant levels of detection sensitivity, pertaining to their unique physical, chemical, mechanical magnetic
and optical features [19]. Several nanomaterials like nanoparticles, nanotubes,
nanowires, nanorods and nanostructured surfaces have been widely explored in the
eld of biological signalling due to their excellent hardiness, portability as well as
electrical and mechanical properties [20]. Some of the fundamental properties of
nanomaterials like tunnelling and quantum effects and high surface-to-volume ratio,
make these nanobiosensors inimitable in today’s eld of disease diagnosis [21].
Therefore, the high selectivity and sensitivity of biosensor ease out early diagnosis and management of diseases through facilitating timely therapy decisions and
implementation of nanobiosensor can improve the assessment of the prognosis of a
disease and its progression important for the effective treatment of many diseases
[22]. This book chapter will help to develop an overall knowledge about recent
development in the eld of nanobiosensor with special reference to their application
in the treatment of human inammatory diseases (Fig.3.1).
3 Recent Advancements in Developing Nanobiosensors for Treating Inammatory…
Fig. 3.1 Recently developed nano-biosensors and their applications
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3.2 Technological Outlines inDeveloping Nanobiosensors
Nanobiosensors are the result of modern advancement in the elds of nanotechnology and electronics fabrication. This new generation of nanoscale biosensors has
taken huge strides in making nanobiotechnology extremely useful for disease diagnosis. Nanobiosensors are very similar in their mode of action to other biosensors
that measure a biochemical event employing optical, electronic or magnetic detection methods except for they use extremely compact probes for transducing the
signal [23–25].
Modern health monitoring and disease detection procedures rely immensely on
rapid, extremely precise and real-time detection of biological events which in turn
aids in rapid decision-making and life-saving manipulations. Nanoscale biosensors
that incorporate nanotechnology with biological detection molecules have helped
scientists achieve this feat very efciently [7, 26].
3.2.1 Importance ofNanotechnology inBiosensing
Nanobiosensors are essentially biosensors which have nanomaterials incorporated
at their core. A nanomaterial comprises of nanoparticles (NPs) that are less than
100nm at least in one dimension [7]. When scaled down to a nanoscale, most materials have most of their constituent atoms located at or near their surface and exhibit
novel properties that cannot be extrapolated from their bulk behaviour [24].
Nanobiosensors show signicant advantages in terms of sensitivity and specicity,
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in contrast to old-school biosensors. From the detection of biomolecules like nucleic
acids, proteins, immunologic molecules to environmental pollutants or any other
small and sparse molecules, the detection prowess of nanobiosensors has established their potential in therapeutic, food and drug quality or environmental assessment [23, 24].
Nanoscale biosensors can overcome many obstacles that impede the widespread
use of afnity biosensors without many of the major drawbacks. For example, in
case of designing afnity-based biosensors, two major targets are to achieve lower
detection limit and to detect multiple analysts for a single sample [7]. Lower detection limit for example can be achieved by altering the thermodynamics of the afnity reaction [7]. Using nanomaterials in developing biosensors provides a surface
area-to-volume ratio far greater than using bulk materials. This, in turn, aids the
nanobiosensors to be extremely sensitive to even trace amounts of target molecules [27].
A. Chakraborty et al.
3.2.2 Classication ofNanomaterials
Nanomaterials can be broadly classied into four types according to size and dimensions namely: zero dimensional (0D), one dimensional (1D) and two dimensional
(2D) [28, 29].
In 0D nanomaterials, all three dimensions of materials are in nanoscale, i.e.
<100 nm. Nanoparticles of metals like silver, gold, palladium, etc. and quantum
dots are amongst the most used 0D nanomaterials.
1D nanomaterials usually have two of the three dimensions in the nanoscale,
while the other in the macroscale. They are mostly lamentous in shape with their
diameter in nanoscale. Metal nanoparticles or quantum dots are often used to
develop these structures that include nanobers, nanowires, nanotubes, etc.
In 2D class of nanomaterials, two dimensions are in macroscale and one dimension is in nanoscale, i.e. they have very low thickness of <100nm while extended in
the other two-dimensional plane. Example includes nanolms, multi-layered nanolms, nanosheets, etc.
Except for these three groups of nanomaterials, most other materials used in
biosensor development are three-dimensional (3D), which have all dimensions in
the macroscale rather than in the nanoscale. However, these bulk materials may be
composed of individual building nanoblocks belonging to the nanoscale.
3.2.3 Nanomaterials Used inDesigning Biosensors
A biosensor is typically made up of three components:
A bioreceptor that serves as a template for the analyte binding.
3 Recent Advancements in Developing Nanobiosensors for Treating Inammatory…
85
A transducer that, as its name suggests, transforms the binding biochemical energy
of analyte with its bioreceptor to electrical energy, and
A detector system that typically receives the electrical signal from transducers and
amplies and documents the signal to a perceivable form.
The basic blueprint for nanobiosensors is exactly the same. In addition to these
features, the transducer portion of a nanobiosensor must have immobilization property that immobilizes bioreceptors on its surface.
Nanoparticles Metal nanoparticles show great potential in terms of stability and
sensitivity at low concentration of analytes, when used in electrochemical nanobiosensors. Noble metal nanoparticles are widely used in biosensors for their exibility
of sizes and compositions that suits different applications. Noble metal nanoparticles have been found to possess exemplary therapeutic efciency in connection with
antimicrobial properties, low doses of treatment and negligible toxicity [30]. Among
the top used nanoparticles, gold nanoparticles when used to immobilize the biocomponents have shown excellent biocompatibility and electroanalytical potential.
Enzyme immobilization using nanoparticles provides greater stability in the biosensors for longer along with improving its analytical sensitivity. According to
Mukherjee etal. (2013), naturally existing enzymes like polyphenol oxidases (PPO)
are found to hold efcacy in wide temperature and pH ranges especially when
immobilized on some matrix [31]. Silver nanoparticles (AgNPs) are a second group
of commonly used nanoparticles in biosensing and drug-delivery systems. An
attempt by Dey etal. (2016) utilized AgNPs as efcient in-cell delivery agents for
antibiotic and antilarial drugs by stabilizing them via a non-toxic supramolecular
hydrogel network, following previous reports of potency of DNA and SHGelcapped AgNPs against various pathogens and parasites. A very interesting example
of green synthesis approaches like harnessing sunlight to generate AgNPs has also
been set forth by the same group [32].
Lanthanide luminescent nanoparticles, having prolonged light-emitting property,
make them excellent probes for nanoscale time-resolved uorometery biosensors.
Iron oxide nanoparticles due to biocompatibility and paramagnetic properties
have excellent immobilization property for the biocomponents [7, 26].
Nanowires Being one-dimensional system, current ow in nanowires/nanobers
is extremely sensitive to minor alterations generated by interaction with biocomponents. Its 1D nature means current ows almost at the surface. Due to these properties and also its inherent nature to bind to biological analytes to their surface helps
in generating direct, label-free electrical nanobiosensor. These biosensors based on
eld effect transistor principles thus are very effective in detecting a host of analytes
ranging from DNA sequences to cancer biomarkers to even whole viruses [7, 26].
Carbon nanotubes (CNT) Carbon nanotubes are other 1D nanomaterials that due
to their unique electrical and mechanical properties have found their way in several
nanobiosensing applications. They can be single-walled or multi-walled CNTs, and
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have high stability, excellent thermal conductivity and high surface-to-volume ratio.
Due to these unique features, even minor disturbances on its surface (e.g. binding of
analytes) alter its electronic conductance and generate a strong signal. Thus, CNTbased nanobiosensors are widely used in healthcare as well as environmental detections [7, 29].
Graphene Graphene, formed by sp2 electron-hybridized carbon atom, is an atomi-
cally thin layer with excellent electron transport properties, high thermal conductivity, adjustable optical property, extremely high tensile strength and superior specic
surface area. Its structure with a porous framework provides excellent opportunity
for surface immobilization of bioreceptors and provides stable surface for bioanalyte interactions. Thus, graphene and graphene derivatives have been extensively
used in developing optical, uorescence, impedance and electrochemical biosensors
that can detect numerous analytes such as biological macromolecules like DNA,
cytochromes, glucose, cholesterol, vitamins, catechol and inorganic molecules
including heavy metals and gases [25, 33].
Quantum dots Quantum dots are modern-day zero-dimensional nanoparticles
having 1–10nm size, which have gained extensive utility in contemporary nanobiosensor designs. They have unique optoelectrical properties including wide excitation spectral range, narrow, sharp and controllable emission spectral band, low
photo-bleaching and superior photochemical stability. The photoemission of quantum dots is related to its size and surface structure and hence, when molecules and
ions bind to their surface, altering their size and surface properties, the emission
intensity or colour changes. Instances have been found of associating different biological heteroatoms with graphene quantum dots to enhance their photoluminescence and electrochemical performance, to eventually conjugate these with
streptomycin, a common antibiotic and successfully develop drug-conjugated biocompatible quantum dots [34].
Multiple biosensing applications that require optical functions that include biosensing of biological macromolecules, pharmaceuticals or other organic analytes,
use quantum dots due to their high sensitivity, size-dependent emission property,
cost-effectiveness and smaller size [35]. Figure3.2 could be referred for overviewing an outline of the different constituents of different nanobiosensors.
3.3 Methodologies Involved inTransduction
Broadly there are two types of biomolecular transduction pathways: label-based
transduction and label-free transduction.
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87
Fig. 3.2 Popular technologies for developing nanobiosensors
3.3.1 Label-Based Biosensors
The native physicochemical properties of most of the biological analytes are often
not enough to be detected by biosensors. Thus, labels such as uorescent, radioactive or enzymatic tags are required to be attached to the target analyte. The signal
thus generated is in fact the proportional reading of the labels tagged with target
analytes.
In case of uorescence biosensors, target analytes (e.g. antigens) with uorophore tag are applied on surface-immobilized detection probes (e.g. antibodies) and
when they bind to the probes, the uorescence is generated [7].
Radioactivity-based detection that uses radioisotopes as labels is a technique of
choice in case of applications requiring high degree of precision and sensitivity.
However, because of the hazardous nature of the radio-labels, the technique is usually restricted to low-throughput applications [36].
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Biosensors in clinical sample testing and cell analysis often use enzymatic tags
for their highly sensitive nature. Among several obstacles of enzymatic labelling
technique, the most disrupting one is its loss of enzymatic activity over time [36].
But the system is by no means awless and comes with certain drawbacks. Apart
from being quite labour-intensive and time consuming, often during the labelling,
purication and multiple wash steps involved in this technique, a substantial risk of
sample loss remains. Moreover, binding of different tags often may block the active
binding sites for the analytes and may result in altered binding properties [7, 36].
A. Chakraborty et al.
3.3.2 Label-Free Biosensors
With astounding progress in the eld of nanotechnology and by help of modern
micro-fabrication processes, label-free detection of biomolecules is now possible
with extreme precision and high sensitivity. The modern nanobiosensors don’t
require ligands but use the intrinsic properties of biomolecules such as molecular
weight, charge, dielectric property, magnetic eld, refractive index, etc. to detect the
presence of analytes. Label-free biosensing provides accurate and fast data about
selectivity and afnity of bioactive molecular interactions often including the information of binding kinetics and thermodynamics [7, 36].
Depending upon the type of transducers used, label-free nanobiosensors can
broadly be divided into optical and non-optical types. The typical non-optical labelfree nanobiosensors again can use different detection methods that include massbased, electrochemical and electrical, acoustic wave, magnetic detection
methods, etc.
Label-free optical biosensors are mostly based on surface-plasmon-resonance
(SPR). There are quantum dot-based biosensors, bio-photonic cell-based biosensors
or optical resonator-based biosensors also gaining popularity among optical labelfree nanobiosensor fabrication. Electrometric biosensors detect the interaction
between analytes and bioreceptors in terms of change in current or voltage and can
include voltametric, amperometric or impedance-measuring transducers. Massbased mechanical detection biosensors on the other hand use nanoscale cantilevers
for detecting change in resonance in free and analyte-bound sensors to determine
mass of the biomolecule [7, 36, 37].
3.4 Different Nanobiosensing Techniques
Biosensors can be categorized according to the basic technologies incorporated in
signal transduction and biorecognition process. The biorecognition element in a
biosensor binds to the target analytes and biological signal is converted to an electrical signal legible to the detector. Here we shall outline some of the most popular
nanobiosensing technologies.
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3.4.1 Optical Sensing
Optical biosensing involves an optical transducer and bioreceptor molecule, where
the optical transducer converts a biological event to electronic events in the presence
of light. The analytes binding with bioreceptor induces a change in amplitude,
phase, polarization, absorption, etc. picked up by the transducer. Modern optical
nanobiosensors also use SPR, Raman scattering and chemiluminescence.
Absorption-based optical sensing Absorption-based optical nanobiosensors
detect the change in light absorption due to the changes in the analyte concentration.
The incident light absorbed by the sample is measured by an optical detector [27].
Surface plasmon resonance-based optical sensing Surface plasmon resonance
(SPR) is a technique that measures surface activity. Plasmons are free electrons
formed at the metal–dielectric interface. Plasmons produce an electric eld known
as an evanescent wave. Polarized monochromatic light striking the prism and solution interface above a certain angle known as resonance angle generates total internal reection. In this situation, the photons resonate with the surface plasmons and
are themselves transformed into plasmons creating a resonance wave, which can be
recorded. Noble metal, e.g. gold (Au), silver (Ag), etc. lms coated over prisms can
be used to create surface plasmons and are widely used for fabrication of biosensors. Any change on the metal surface due to binding of biomolecules, can alter the
momentum changing the resonance angle for SPR resulting in a SPR shift. This
shift can be recorded and interpreted in terms of biomolecular interaction, presence
or activity. Noble metal nanoparticles are used in nanobiosensors instead of thin
lms which are able to amplify SPR signals to many-folds providing ultrasensitive
detection [7].
Fluorescence-based optical sensing In any uorescence detection mechanism, an
excitation light source, uorophore molecules, lters isolating emission spectra
from excitation spectra and a detector are the core components. Energy is provided
by the light source and absorbed by the uorophore, producing an excited state. The
most modern and commonly used uorescence detection system in nanobiosensing
uses uorescence resonance energy transfer (FRET), a non-radiative quantummechanical technique. FRET using non-radiative mode of energy transfer among
nearby uorophore/chromophores is responsible for efcient energy transfer, making them extremely sensitive to biomolecular interactions. FRET-based nanobiosensor is a combination of a bioreceptor attached to a pair of uorophores mostly
quantum dots fused with bioreceptors. Graphene-based quantum dots with gold
NPs for DNA detection is a very good example for the same. Gold nanoparticles are
also used as efcient universal uorescence quenchers in nanobiosensing. When the
target molecule (mostly DNA) interacts with the bioreceptors immobilized with this
system, the conformational change restores the uorescence which is then detected.
Chemiluminescence is a similar process to uorescence except that it utilizes
energy from chemical reactions to generate excitation energy [20, 27].
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3.4.2 Electrochemical/Electrical Sensing
The biosensors based on these technologies perform biorecognition of analytes primarily by recording the change in electrical events like current, voltage, impedance,
etc. Electrochemical biosensors perform label-free detection of analytes with high
throughput by interpreting electrochemical reactions between bioreceptors and analytes on the transducer surface. Based on the operating transduction principle, the
electrical sensing mechanism can be of the following types [27, 38].
Potentiometric/voltametric sensor: while applying a constant current, biochemical
reactions between bioreceptors of a biosensor and the analytes generate change
in potential which is measured by the voltametric biosensors. Voltametric nano-
biosensors use eld-effect transistors (FETs) of different kinds like ion-sensitive
FETs or graphene FETs, where biomolecular immobilization is achieved using
carbon nanomaterials like CNT or graphene and the signal is measured by accu-
mulation of biomolecules on the interface [38].
Amperometric sensor: In the presence of a constant potential, chemical interaction
between the analyte and bioreceptor immobilized on the transducer surface gen-
erates change in current. This is measured by amperometric biosensors. The
electrode of an amperometric nanobiosensor is usually made from a noble metal
nanomaterial, of carbon nanostructures covered by immobilized bioreceptors on
their surface. On application of a constant potential, the generated current by
catalytic conversion of biomolecules at the surface is measured [27].
Conductometric biosensors: they measure small change in the conductance in the
analyte solution due to their interactions with bioreceptors. Conductometric
transducers are extremely small two-electrode systems that, when a thin electro-
lyte layer is applied, measure the conductivity of the solution [38].
Impedimetric sensor: electrical impedance is the amount of resistance that a circuit
imparts on the ow of current at a certain applied potential. Any bioanalyte when
reacts with surface-immobilized bioreceptors the resistance is altered. This alter-
ation in impedance at the analyte-receptor interface is detected by the transduc-
ers to generate a reading. Single-wall CNTs, noble metal nanoparticles, etc. are
very important nanomaterials in the construction of this kind of transducers [38].
3.4.3 Magnetic Sensing
Specially designed ferrite-based magnetic nanoparticles are used for designing
magnetic nanobiosensor. The magnetic nanoparticles are fabricated by incorporating iron with transition metals forming an alloy having unpaired electrons in their
outer orbital making their magnetic property exible to various use. Magnetic
nanoparticles are great tools to separate and enrich analyte sample that are magnetically labelled. More recent techniques involve superconducting quantum
3 Recent Advancements in Developing Nanobiosensors for Treating Inammatory…
interference devices (SQUID) that use superparamagnetic nanoparticles to rapidly
detect the analytes in most cases antigens using specic corresponding antibodies
tagged with magnetic nanoparticles [20, 24].
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3.4.4 Mass-Based Sensing
Mass-based mechanical detection of analytes is achieved by the use of nanoscaled
cantilever sensors. Nanocantilevers are exible beams, xed at one side and are usually made up of silicon or quartz. Biorecognition molecules are immobilized on the
cantilevers that bind target molecules if present in the analysis media. This added
mass of the analytes alters the conformation or resonating properties of the nanocantilevers. Being in nanoscale increases the sensitivity of the sensor as smallest
addition of mass will affect those properties signicantly. Also, one of the inherent
properties of any nanomaterial being its increased aspect ratio provides more surface for biomolecules to attach on the nanocantilevers [26].
Depending on the mode of excitation, nanocantilever biosensors can be
deection- based or static nanocantilever sensor and resonance-based or dynamic
nanocantilever sensor.
In static cantilevers, binding of analytes deects the beam from its original position in proportion to the mass-binding. In dynamic sensors, the captured analyte
will add mass to the cantilever to affect its resonating frequency. In both cases,
nanocantilevers that are not bound with analytes are used as controls to compare the
shift in the conformation or resonance [7, 26].
3.5 Tribology ofNanoparticles intheContext
ofDeveloping Nanobiosensors
Tribology, the study of interacting surfaces in relative motion, plays a pivotal role in
optimizing the performance, reliability and durability of nanobiosensors. Tribology
focuses on the friction, wear and lubrication between interacting surfaces [39].
Friction, wear and lubrication between materials in contact are of utmost importance in scientic applications [40]. Tribological considerations are critical in nanobiosensors as they directly inuence the functionality, sensitivity and lifespan of
these sensors. Nanomaterials, such as nanoparticles, nanowires and nanostructured
lms, are commonly employed in the fabrication of nanobiosensors due to their
unique physical, chemical and biological properties. These materials exhibit distinct
tribological behaviours that signicantly impact the performance of nanobiosensors
in disease detection applications. Highly sensitive nanobiosensors are manufactured
by incorporating nanomaterials designed with nanoelectromechanical systems
(NEMS) that give complex tribological properties. With the implementation of