Nano-Scaled Materials and Polymer Integration in Biosensing Tools: History
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Contributor:
Hichem Moulahoum
,
,
Emine Celik
,
Suna Timur

The evolution of biosensors and diagnostic devices has been thriving in its ability to provide reliable tools with simplified operation steps. These evolutions have paved the way for further advances in sensing materials, strategies, and device structures. Polymeric composite materials can be formed into nanostructures and networks of different types, including hydrogels, vesicles, dendrimers, molecularly imprinted polymers (MIP), etc. Due to their biocompatibility, flexibility, and low prices, they are promising tools for future lab-on-chip devices as both manufacturing materials and immobilization surfaces. Polymers can also allow the construction of scaffold materials and 3D structures that further elevate the sensing capabilities of traditional 2D biosensors.

  • nanocomposites
  • polymer scaffolds
  • nanoparticles
  • optical sensing

1. Polymer and Biopolymer Nanocomposites

Polymers have been considered prominent candidates for creating an ideal matrix for entrapment and immobilization of biomolecules in the analytical sciences. Characteristics of most polymers, such as high conductivity, ease of biofunctionalization, flexibility, biocompatibility, highly modifiable chemical functions, etc., make them attractive for biosensor development in different fields from environmental analysis to biomedical applications [1]. The experience with polymeric and co-polymeric materials also demonstrates the successful application of functional polymers for the immobilization of enzymes [2], micro-organisms [3], antibodies [4], and aptamers [5] in the design of electrochemical biosensors. Historically, polymers have been seen as a practical material choice for electrochemical devices, but recent advances in nanotechnology and the creation of nano-scaled materials has allowed for even further evolution in the biosensor field development due to new intrinsic optical, electrical, and mechanical properties.
Efficient immobilization of bioreceptors (or other components) and optimal signal transduction are crucial for biosensors. Polymers are still key coating matrices for nanomaterials through the fusion of nano-objects and polymers. These combinations have led to the emergence of new nano-scaled hybrid materials or polymer nanocomposites. They have been further employed to construct polymer nanocomposite-based biosensors to obtain highly sensitive and reliable analytical devices by improved catalytical and chemical reactivity, surface specificity, enhanced electrode kinetics, controllable synthesis and morphologies, higher stability, and biocompatibility [6]. As an alternative to synthetic polymers, some features of biopolymers, such as natural origin, biodegradability, recyclability, lower antigenicity, and their suitable interaction with living systems, make them powerful tools for biosensor fabrication [7][8].
Polymeric and biopolymeric nanocomposites refer to a hybrid structure in which a polymer matrix is used as a substrate, and nano-scaled organic or inorganic materials are used as fillers. Typically, the polymers of poly(lactic acid) (PLA), poly(ethylene oxide) (PEO), poly(lactic-co-glycolide) (PLG), poly(N-isopropyl acrylamide) (PNIPAM), and polyurethanes, etc. have been utilized as the matrix phase of polymeric nanocomposites. Biopolymeric nanocomposites, which are also called “Bio-nanocomposites”, “bio-hybrids”, and “green nanocomposites” by the popular terms of recent years, are made up of a nanosized additive in naturally occurring polymers including cellulose, chitin, collagen, silk, keratin, alginate, lignin, starch, polyhydroxyalkanoates (PHA), etc. [9]. The merging of nanosized filler materials into the polymeric matrix produces interesting and improved mechanical, thermal and optical properties. According to this reinforcement strategy, it can be said that filler materials act as molecular bridges enhancing and controlling dimensional stability, flexibility, strength, toughness, durability, thermal stability and conductivity, optical properties (color and transparency), size, distribution, and shape [7][10][11]. Organic materials (carbon nanotubes (CNTs) and graphene) and inorganic materials (silicates and metal/metal oxides) are the kinds of nanofillers used to prepare nanocomposites made of polymers [10][11]. The characteristics of polymeric nanocomposites are affected by choice of both the filler and matrix. For instance, while the type of polymer matrix significantly determines the hydrophobicity, transparency, strength, toughness, controlled ionizability, crystallinity, functionality, biocompatibility, and biodegradability, the choice of filler considerably affects the structural and functional properties. Hence, unique polymer nanocomposites can be synthesized by various combinations of nanofillers. This diversity provides application-oriented strategies via selection of filler nano-objects for the desired properties of specific fields, including medicine, diagnostics, biomedical applications, food packaging, optoelectronic devices, biosensing, bioimaging, tissue engineering, cosmetics, energy, etc. [9][12].
In parallel to their flexible functionalities and fascinating properties, polymeric nanocomposites have been extensively studied to improve sensor performance and have remarkably allowed the fabrication of many novel biosensors in recent years [13][14][15]. For example, while quantum dots–polymeric nanocomposites exhibit excellent fluorescence properties that can be used in optical biosensors, CNTs–polymeric nanocomposites provide significant enhancement in the mechanical property that can be adapted to an optoelectronic sensing device [12]. The utilization of polymeric nanocomposites provides needs-based designs. It brings additional key performance parameters, including higher sensitivity and selectivity, lower detection limits, good reproducibility, and stability by providing a large and easily adjustable surface area, higher electrical conductivity, and fast electron transfer rate [15].
For polymeric nanocomposites, chemists and material scientists have described various synthesis methods, including ion exchange, template synthesis, sol-gel, in-situ polymerization, hydrothermal route, melt intercalation techniques, etc. [12][16]. The successful design of a polymeric nanocomposite with any required property is a critical step toward the control of interfacial interactions between the nanofiller and the polymer matrix. Understanding the influence of the filler on the size, shape, orientation, dispersion, and compatibility of the polymer matrix is the most important consideration. When creating a new polymer nanocomposite material, an effective formulation is required by considering three main approaches: rationality-based design, functionality-based design, and tailored property-based design. In the design of polymeric nanocomposites, process route, temperature, pressure, and time are the parameters required to be controlled during processing. The nanofiller choice needs to consider the filler shape, size, type, volume, weight, and orientation. In contrast, the matrix preparation needs to consider the kind of polymer, surface nature, and chemistry. Based on the above, the combination of the nanofiller and the polymer matrix must be achieved at a nanoscale level with chemical compatibility and homogenous dispersion.
Polymer nanocomposites are microstructures of hybrid organic–inorganic materials that can be formed into three types; unintercalated (or microcomposite), intercalated (and/or flocculated), or exfoliated (or delaminated). These microstructural forms are controlled by the synthesis method. Of these various synthesis methods, the melt-blending method is eco-friendly because of the lack of solvent usage and is industrially scalable due to its cost effectiveness, however the need for a high temperature that can damage the surface of nanofiller is its main disadvantage. On the other hand, the in-situ polymerization technique provides better exfoliation in comparison to the melt intercalation method. In the case of sol-gel technology, disadvantages such as high temperature, which can cause the degradation and aggregation of polymers, make it uncommon. There are different synthetic methodologies including organic treatment and chemical modifications for the polymer nanocomposites manufacturing process [17]. Click chemistry and ring opening of epoxides and aziridines are very efficient and common chemical concepts in the fabrication of polymer nanocomposites. In particular, click reactions are versatile coupling methods due to advantages that include methodological simplicity, high reaction yield and high reaction rates, moderate reaction conditions, easily removable byproducts etc. CuAAC click reaction, metal-free click reaction, Diels–alder reaction, and Thiol-ene and thiol-yne reactions are the commonly employed click reactions in the fabrication of polymer nanocomposites [18]. Additionally, different surface modification strategies have been developed, something which is also a very critical step in biosensing chemistry and design. Surface modification techniques significantly impact the nanocomposites’ structural and functional properties such as reactivity/chemical reactivity, biocompatibility/bioactivity, hydrophobicity, surface energy, dispersion/stability, and surface roughness. Surface modification can be achieved through different reactions with coupling agents and/or surface adsorption and polymeric molecules’ covalent or non-covalent bonding-based grafting. Along with such functionalization techniques, the reported polymer-nanocomposites-modified electrodes are very promising tools to enhance sensing capabilities in terms of high sensitivity and good selectivity for different types of targets such as drugs, heavy metals, pesticides, pathogens, etc. Polymer nanocomposites still hold a great strength in biosensor design since they provide variable morphologies and architectures on electrode surfaces such as films, vesicles, and dendritic structures [12]. While choosing the nanocomposites-based structural design, the main strategy is to add a nanofiller according to the need to give the final sensor system a targeted feature such as magnetism, fluorescence, electroconductivity, strength etc. The characteristics of a nanofiller considerably affects the properties of the polymer nanocomposites. The incorporation of a nano-scaled structure can add a new feature or improve an existing property. In order to obtain a nanocomposite structure with magnetic properties, magnetic beads can be added to the composite structure, on the other hand quantum dots can be used as nano-modifier to prepare fluorescent polymeric nanocomposites. For the strength enhancement and hydrolytic stability, silica nanoparticles are a very appropriate choice and for their excellent mechanical stability, CNTs are very attractive nanofillers for the polymer nanocomposites. These unique properties, which are gained by adding nanofillers to polymer nanocomposites, significantly increase the analytical performance of the fabricated sensors. Table 1 represents the common nanofillers and their effect on polymer nanocomposite and advantages for the final properties of the fabricated biosensors.
Table 1. The effect of several nanofillers on the improved properties of polymeric nanocomposites and their sensors.
Nanofiller Polymeric Composite Effect for the Fabricated Composites Advantages of the Sensors Ref.
Nanoclay OMMT/PLA Improved thermal and mechanical property Improved surface morphology and surface reflectance, modified optical properties [19]
Graphene GC-COOH Electroactivity High electroactivity and easy assembly, high sensitivity, [20]
CNT Chitosan modified by ferrocene
and CNT		 Increased surface area and decreased effective distance between mediator molecules Increased recorded analytical signal, and measurement sensitivity [21]
PAMAM dendrimer PAMAM-PPy Functionality and increased quantity and homogenous distribution of attached biomolecules Efficient electron transfer, reversible redox system, and simple reaction procedure [22]
Oleic acid-modified MNPs Magnetic cyclodextrin vesicles Magnetic property Higher sensitivity [23]
Nano rod and Quantum Dot TiO2 Nanorod/TiO2 Quantum Dot/Polydopamine Strong light absorption and excellent
photocatalytic activity		 Stronger photoelectric response under visible light [24]
PLA: Polylactide, OMMT: Organically modified montmorillonite, GC-COOH: Carboxylated chitosan-functionalized nitrogen-containing graphene, CNT: Carbon nanotube, PAMAM-PY: poly(amidoamine) dendrimers and polypyrrole film, MNPs: Magnetic nanoparticles.

2. Conducting Polymer Nanocomposites

The four valance electrons of some polymers’ constructive carbon atoms are not fully used up in covalent bonds. These polymers are well known as conjugated polymers in which the electron delocalization provides high charge mobility along their carbon backbones. The conjugated polymers can possess semiconducting features or metallic properties depending on the number and kind of atoms within the repeated polymeric units. Conjugated polymers can also be transformed into conducting polymers by doping processes that change the number of π-electrons [25]. Heeger et al. received the Nobel Prize in Chemistry for the discovery of the first conducting polymer (polyacetylene). Conducting polymers (CPs) exhibit remarkable features such as high mechanical, electronic, optical, and environmental stability and low operating temperature and are lightweight, offer a simple synthesis, and economical behavior [26]. These outstanding features have led to the fabrication of optical wires, gadgets, and biosensor devices, such as sensor chips for diagnostic and environmental monitoring purposes [27][28].
The formation of composite and nanocomposite by the addition of fillers has been widely recommended to enhance the physical and chemical features of CPs. A composite typically consists of two or more constituents in which each component carries its features to the final structural material. Using nanomaterials of different types and shapes as a reinforcing phase in the CPs matrix phase creates a conducting polymer nanocomposite (CPNC). Several methods to produce CPNCs include electrochemical encapsulation, colloidal dispersions, in situ polymerization with nanoparticles, and coating of inorganic polymers [29]. The properties of CPNCs can be tuned by varying the matrix and filler, which results in millions of combinations usable in different applications. Alternative carbon nanomaterial (CNMs) fillers such as single-walled and multi-walled carbon nanotubes, fullerenes, carbon nanofibers, nanospheres, and graphene have been extensively used for CPNCs preparation [13]. The unique features of carbon nanomaterials, such as their environmental stability, surface area, and other properties (physical, chemical, thermal, and electrical), make them unique materials for the twenty-first century. Substantial efforts have been allocated to produce CPNCs with superior fundamental and technological assets through CNM and CP combination [30].
Other than CNMs, metal nanoparticles (silver, gold, platinum, etc.) and their oxide forms have been employed to create CPNCs with advanced features due to their different compositions and dimensions [31][32]. Various techniques such as electrochemical or chemical methods [33], sonochemical methods [34], sol-gel techniques [35], ultrasonic irradiation [36], and photochemical preparation [37] have been actively used to incorporate metals or metal oxide fillers into the preparation of conducting polymer nanocomposites.
As an active biosensing platform, CPNCs such as nanocomposites of polypyrrole (PPy) and polyaniline (PANI) conducting polymers have shown high biocompatibility with cells and biological tissues. These features influenced researchers to employ CPNCs in tissue engineering, bio-electrodes, drug delivery, and biosensors to detect biological and synthetic moieties [38].

3. Molecularly Imprinted Polymer Nanocomposites

Molecular imprinting technology is of great interest in biomimetic molecular recognition. Molecularly imprinted polymers (MIP), in which a target-specific cavity is created using a template, are considered an essential alternative to natural antibodies in bioanalytical devices. Due to their low cost, flexibility, outstanding chemical stability, and high recognition ability, MIPs have been used to fabricate biosensors in numerous studies. These artificial receptors suffer from some drawbacks, including long response time, heterogeneous structure of binding cavity, diffusion rate, etc. The main reason for these limitations is slow binding kinetics arising from the bulky forms of MIPs such as monoliths, thin films, microspheres, etc. The binding efficiency decreases as the recognition sites remain inside the MIP structure. Also, the low surface area of a general MIP results in a limited amount of recognition sites. Low affinity caused by these problems in biosensing systems has been the main focus for transforming the binding event into a successful signal [39]. Hybrid approaches to obtain MIP nanocomposites have great perspectives on enhancing biosensing performance by overcoming the limitations of traditional imprinted polymers. In the hybrid-material strategy, a variety of functional nanomaterials could be used. For their synthesis, the thickness of MIP films is controlled by an inorganic core. Some formation methodologies of such core–shell MIP nanocomposites are controlled through living radical polymerization (CRP, reversible/addition/fragmentation, chain transfer polymerization (RAFT), or atom transfer radical polymerization (ATRP) [40]. The application of molecular imprinting technology in nanocomposites combines the unique advantages of MIPs, such as affinity, physical and chemical stability, low cost, etc., with optical and/or electrical properties of nano-scaled materials. This synergy makes MIP nanocomposites very feasible tools for biosensor construction. Since the amount of high affinity imprinted sites alone is not enough for the sensitivity of a biosensor, the enlargement of surface area and manipulating interfacial properties by nanomaterials are important approaches in sensing strategies. Additionally, molecular geometry is a crucial factor to improve the binding ability of MIPs. Hence the sensitivity of a biosensor is also directly controlled by the imprinted polymer nanocomposites [41]. To increase detection selectivity and sensitivity, CNTs, AuNP, graphene, QDs and, SiO2, noble metal, Fe3O4 nanoparticles are the common modifiers of MIPs [42][43]. According to the target-oriented sensing strategy, the expected function can be achieved by choosing one of these nano-modifiers to develop different analytical methods, including optical, fluorescence, and electrochemical. For instance, among the nanomaterial-MIP hybrid materials, QDs are typical nanostructures used in fluorescence-based sensing applications due to their photostability and size-dependent fluorescence spectra. Although different nanofluorophores can be used in MIP fluorescence sensors, QDs have received greater attention because they offer narrower emission and broader absorption spectra. According to reports, there are several developed QD–MIP-based sensing methods [42][44]. In addition to fluorescence property, magnetic features can be added to MIPs for the construction of electrochemical biosensors. For example, Fe3O4 nanoparticles has been reported as an appropriate candidate by providing easy and quick fabrication technology to a biosensor system. Thanks to their excellent properties such as stability, catalytic activity, non-toxicity, and high surface area, Fe3O4 nanoparticles have been utilized to improve the sensitivity and selectivity of biosensors. Hence, Fe3O4 nanoparticles provide increased sensitivity while MIP has a unique cavity for the target molecule, and this magnetic MIP nanocomposite creates a precise sensor surface [45]. Another idea for MIP-hybrid-based fabrication strategies has been to utilize graphene for better electron transfer, higher mechanical strength, and increased specific surface area. This approach improves electrochemical assays’ performance due to the graphene nanohybrid-based electrochemical signal amplification [41]. Similarly, to add a particular function to a biosensing surface, SiO2 nanobeads can be used to form nanolayers for enhanced sensing ability [46]; TiO2 nanoparticles to provide high surface area, improved adsorption of the target, and quick electrochemical response [47]; silver nanoparticles (AgNPs) to incorporate optical properties [48]; and MWCNTs to accelerate the electron transfer [39] could be developed.

4. Hydrogel Nanocomposites

The 3D polymeric networks in hydrogels can infuse a significant quantity of water and soluble molecules [49]. The weak mechanical strength of hydrogels is a considerable drawback limiting their performance where strength, elasticity, and endurance are highly demanded [50]. Other physicochemical criteria (diffusion, swelling, functional groups) should be closely considered when selecting materials for hydrogel manufacturing. Recent approaches in hydrogel optimization have led to the development of new hydrogel varieties such as nanocomposite hydrogels [51] and double network hydrogels [52]. The nanocomposite hydrogels containing various physically/chemically crosslinked nano-scaled structures among polymeric chains have shown novel properties and behaviors. Nanocomposite hydrogels can be created using different nanomaterials, including carbon-based nanomaterials, polymer NPs, inorganic/ceramic NPs, and metal NPs [53].
Creating nano/micro-sized matrices that benefit the unique properties of both hydrogels and nanomaterials is the main challenge in developing nanocomposite hydrogels. Typically, hydrogels’ flexible and 3D polymeric configuration can host different types of materials as a “guest” [54]. The gelation procedure of the final composite structure happens in any water-based or organic solution resulting in either hydrogel for aqueous media or organogel if made in organic media. Various natural polymers such as chitosan (CS) [55], cellulose [56], alginate [57], collagen [58], and lignin [59], as well as synthetic polymers, including poly(ethylene glycol) (PEG) [60], poly (N-isopropyl acrylamide) (PNIPAM) [61], poly(vinyl imidazole) [62], poly(vinyl alcohol) (PVA) [63], and poly(acrylic acid) (PAA) [64] show the ability to create hydrogels. The “host-guest” interaction between hydrogels and nanomaterials is formed through covalent and non-covalent bonding, such as hydrogen bonding, van der Walls forces, and electrostatic interactions [65]. The application of functionalized nanomaterials can significantly enhance the features of the final nanocomposite hydrogel, namely the mechanical properties and bioactivity [66]. On the contrary, by taking advantage of their chemical structures and forming π-π stacking interactions, non-functionalized nanomaterials (graphene and carbon materials) can also enhance some aspects of hydrogels compared with pure ones [54].
Other than organic-based nanomaterials, inorganic nanomaterials (nanoclays, ceramics, bioactive glass, metallic NPs) are actively utilized to produce nanocomposite hydrogels [53]. The implication of metal NPs and their oxide forms in the hydrogel structure has brought up attractive attributes such as magnetism, electrical and thermal conductivity, and antimicrobial activities. This makes these nanocomposite hydrogels great alternatives as sensors and conductive scaffolds in addition to other applications such as drug delivery [53].
A limited number of inorganic nano-scaled materials can be used directly in sensing platform formation. Inorganic components have specific characteristics, making them very competitive to reach a defined purpose and function. Therefore, these materials are generally used as primary or secondary components. On the contrary, inorganic nanomaterials can be widely utilized as additives to enhance the analytical performance of sensors. Forming homogeneous dispersions of inorganic materials is hard to achieve, whereas using a strong mixing procedure could denaturalize the hydrogel structure. If inorganic additives are effectively dispersed into hydrogels, the obtained sensor can exhibit steady signal transduction and stable performances. Among the wide range of inorganic materials used to improve performance, nano-scaled silica [67], titanium oxide [68], quantum dots [69], and organosilicates [70] are preferred for the functionalization of hydrogels.
Other inorganic nanomaterials such as QDs [71][72][73], noble metal [69][74], and magnetic nanoparticles [75] took part in many research areas. Cadmium Selenide QDs nanocrystals were combined with PEG-based hydrogels and successfully applied for phenol detection [71]. Enzyme encapsulated cadmium telluride QD-based hydrogels with a biocatalysis unit and a fluorescence signaling unit was utilized as a multifunctional material to develop optical biosensors [72]. Magnetic nanoparticles in sensors and biosensors are in high demand due to the limited accessible number of candidate materials.

This entry is adapted from the peer-reviewed paper 10.3390/bios12050301

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