Biosensors research is a fast growing field in which tens of thousands of papers have been published over the years, and the industry is now worth billions of dollars. The biosensor products have found their applications in numerous industries including food and beverages, agricultural, environmental, medical diagnostics, and pharmaceutical industries and many more. Even though numerous biosensors have been developed for detection of proteins, peptides, enzymes, and numerous other biomolecules for diverse applications, their applications in tissue engineering have remained limited. In recent years, there has been a growing interest in application of novel biosensors in cell culture and tissue engineering, for example, real-time detection of small molecules such as glucose, lactose, and H2O2 as well as serum proteins of large molecular size, such as albumin and alpha-fetoprotein, and inflammatory cytokines, such as IFN-g and TNF-
Biosensors have gained enormous attention in recent years in medicine and nanotechnology, and there is a growing interest in its application in tissue engineering. Since the development of the first oxygen biosensor by Lel and Clark in 1962 [
Recently, biosensors have shown immense potential for applications in tissue engineering and regenerative medicine. Both tissue engineering and regenerative medicine are rapidly growing fields in biomedical engineering presenting enormous potential for development of engineered tissue constructs for restoring the lost functions of diseased or damaged tissues and organs [
To precisely sense the biological signals in a cellular microenvironment, a probe with micro- or nano-dimensions is desirable. For this purpose, sensors with nanoscale dimensions, such as nanotubes or nanowires, have been developed for effective biosensing and diagnostics purposes. They can be used to measure pH or functionalized with specific capture molecules to identify very low quantities of biological and chemical species [
In fact, research is in progress to use nanobiosensors in combination with signaling and therapeutic delivery devices for
A biosensor can be defined as “a self-contained analytical device that combines a biological component with a physicochemical component for the detection of an analyte of biological importance.” It is typically comprised of three fundamental components, such as (a) a detector to detect the stimulus, (b) a transducer to convert the stimulus to output signal, and (c) a signal processing system to process the output and present it in an appropriate form Figure
Schematic representation of the working principle of biosensors: (a) interaction between tissue, interphase, and biosensors. Figure
Biosensors can be classified into different types either based on their sensing components or the transducer components as described below.
The biosensing components of biosensors can be divided into two types, namely, catalytic type and affinity type. The catalytic type sensors include enzymes, microbes, organelles, cells, or tissues, while the affinity type includes antibodies, receptors, and nucleic acids. Some of the important ones among these types are discussed below.
The enzymes used as bioreceptor components in biosensors are usually proteins of oxidase type that can selectively react with specific analytes, consume dissolved O2, and produce H2O2 that is an easily detectable compound. Other mechanisms of enzyme based biosensing include the detection of enzyme activation or inhibition by the analyte and the modification of the enzyme properties by the analyte. The enzyme molecules can be directly immobilized on the transducer surfaces using entrapment in gels, attachment through covalent bonding, physical adsorption on the surfaces, or other available techniques [
The use of microbes has a number of advantages as biological sensing component in the production of biosensors. They are present all over and have a great capacity to acclimatize to undesirable conditions and to develop the ability to metabolize new molecules with time. Microbial cells are cheaper than enzymes or antibodies. They can carry out several complex reactions while maintaining their stability. Whole cells can be used either in a viable or nonviable form. Viable cells have gained importance in the manufacture of biosensors and these cells metabolize various organic compounds either anaerobically or aerobically resulting in various end products like ammonia, carbon dioxide, acids, and so forth that can be monitored using a variety of transducers. The use of microbial biosensors is common in environmental fields that include the detection of harmful bacteria or pesticides in air, water, or food and biological oxygen demand [
The compartments located inside the cells are known as organelles. Each of the organelles has individual functions such as lysosome, chloroplast, and mitochondria. Mitochondria are responsible for calcium metabolism and controlling the calcium dependent pathways in cells. Previous studies proved that presence of high concentration of calcium stimulates the mitochondria to open the calcium channels. This bioinspired strategy can be used to measure calcium concentration in medium. Application of mitochondria for water pollution detection is another application of organelles in biosensor [
Cells have been often used in bioreceptors because they have high sensitivity to adjacent environment. The attachment on the surface is the main characteristic of cells, so they can be easily immobilized. They are frequently used to detect global parameter like stress condition, toxicity, and organic derivatives and to monitor the treatment effect of drugs. Cells were also used in ion selective transducers [
Antibodies are proteins produced by B-Lymphocytes in response to antigenic stimulation. The antibody based sensors are also known as immunosensors. Usually, antibodies are used in surface plasmon resonance (SPR) biosensors to design target specific sensors for detecting specific biomolecules. This is a simple mechanism that works through antigen-antibody interaction process. The antibodies are usually linked to the surface of transducers through covalent bonds such as amide, ester, or thiol bonds. The transducer surface needs to be modified by polymers or monomers to introduce functional groups such as carboxyl, amino, aldehyde, or sulfhydryl groups to facilitate conjugation between the antibody and transducer. To date, many antibodies have been made available in the market and used in immunoassays. They are more accurate and faster compared to the traditional assays [
Oligonucleotides integrate in nucleic acid biosensor with a signal transducer. Oligonucleotide probe is immobilized on the transducer to detect DNA/RNA fragments. The detection process is based on the code of complementary nucleotide base pairing, adenine (A): thymine (T) and cytosine (C): guanine (G) in DNA. The hybridization probes in the sensor can then base pair with the target sequences and create an optical signal [
The transducer component of biosensors can be grouped into different types such as electrochemical, optical, acoustic, and calorimetric types.
Electrochemical biosensors are mainly used for the detection of hybridized DNA, glucose concentration, and so forth. Electrochemical biosensors can be classified based on measurement of electrical parameters such as: (i) conductometric, (ii) amperometric, and (iii) potentiometric types. Electrochemical biosensors usually contain three electrodes: a reference electrode, a working electrode, and a counter electrode. The reaction for target analyte takes place on the active electrode surface. The reaction causes either electron transfer across the double layer or can contribute to the double layer potential. These kinds of biosensors are often made by screen printing the electrode patterns on a plastic substrate, coated with a conducting polymer and then some protein is attached. All biosensors usually involve minimal sample preparation as the biological sensing component is highly selective and the signal is produced by electrochemical and physical changes in the conducting polymer layer.
Optical biosensors are usually made based on optical diffraction. These sensors can detect microscopic changes when cells bind to receptors immobilized on the transducer surface. They use the changes in mass, concentration, or number of molecules to direct changes in characteristics of light. Researchers have used optical techniques such as SPR and ellipsometry for the detection of bacterial pathogens [
The acoustic transducers used in biosensors are based on either the bulk acoustic wave or the surface acoustic wave. The transduction is through detection of changes in their physiochemical properties, such as mass density, elasticity, viscoelasticity, or electrical conductivity [
Biosensors can be of immense importance in tissue engineering applications, particularly in maintaining three-dimensional cell cultures [
In clinical applications, biosensor-based monitoring of blood glucose concentration has now become a major diagnostic method to accurately trace diabetes with high levels of glycated hemoglobin (HbA1c). In tissue engineering applications, however, continuous monitoring of glucose in culture media is used as an indicator of metabolic activities of cells [
Accurate and reliable measurement of H2O2 is of paramount importance in both tissue engineering and clinical applications. The monitoring of H2O2 allows detecting the presence of oxidative stress or hypoxic conditions in the cell and tissue culture. Currently, available analytical methods of H2O2 measurement include techniques such as electrochemistry, photometry, and titration [
Extracellular adenosine diphosphate (ADP) and adenosine triphosphate (ATP) are vital multifunctional molecules present in blood, heart, and liver. Apart from its main role in cellular metabolism, ATP is now recognized as an important extracellular signaling agent. It can modulate a number of physiological pathways by activating specific plasma membrane receptors. Luciferase-based methods have long been used for measuring adenosines; however, they have limited application
It is important to measure the activities of functional protein molecules such as bioenzymes released from cells, under different microenvironmental conditions to understand the fundamentals of cell biology for therapeutic, diagnostic, and tissue engineering applications. Matrix metalloproteinase (MMPs), a member of proteinases family, is released by cells as a biological response to their natural tissue remodeling processes [
Biosensors have been used for early detection of cancer biomarkers from blood samples in a noninvasive manner. Surface plasmon resonance (SPR) and electrochemical biosensors have been successfully used for the detection of carcinoembryonic antigen (CEA) biomarkers for early diagnosis of lung cancer in serum [
Different types of biosensors have been developed for detection of pathogenic microbes. Amperometric biosensors have been developed for indirect detection of
Semiconductor quantum dots (Qdots) are one of the most promising optical imaging agents for
Schematics for some recent advancement in biosensors applicable in tissue engineering. (a) Variation of color in quantum dots (blue, green, yellow, and red) based on their emission wavelength. (b) Carbon nanotube based biosensor for detecting various cell secreted biomolecules from tiny amount of sample. (c) Some MEMS based biosensors: (i) SPR: surface-plasmon resonance; SMR: suspended microchannel resonator; NW: nanowire; LFA: lateral flow assay; MRR: microring resonator; QCM: quartz crystal microbalance; BBA: biobarcode amplification assay; IFA: immunofluorescent assay; MC: microcantilever. (ii) static-mode surface-stress sensing by a MEMS device (iii) scanning electron micrograph of dynamic mode MEMS device and (iv) suspended microchannel resonator (SMR). (d) (i) Graphene and its derivatives (graphene oxide, graphene quantum dots) based sensors. (ii) Vertically-oriented graphene based field effect transistor-sensor by direct growth of VG between the drain and the source electrodes. (c) and (d) (ii) reproduced from [
The unique chemical and physical properties of carbon nanotubes have introduced many new and improved sensing devices. Early cancer detection in
The growing need for miniaturization of biosensors has resulted in increased interests in microelectromechanical systems (MEMS) [
Graphene based biosensors have attracted significant scientific and technological interests due to the outstanding characteristics of graphene, such as low production cost, large specific surface area, good biocompatibility, high electrical conductivity, and excellent electrochemical stability [
Graphene derivatives, especially 0D graphene quantum dots (Gdots), are photoluminescent materials derived from graphene or carbon fibers [
The glucose biosensors, as mentioned in earlier section, can be used in tissue engineering for continuous measurements of metabolic activities of cells. Graphene oxide (GO), the precursor material of graphene, has been used as a novel highly efficient enzyme electrode for the detection of glucose in phosphate buffer saline solution (PBS) [
Graphene-based nanocomposite materials have been extensively used in the fabrication of glucose biosensor [
The long-term stability of the developed biosensor was examined over 30 days and was found to be stable after the immobilization of the electrode with GOD. Wu et al. [
Schematic illustration for the preparation of SPEEK functionalized graphene and the biochemical reaction mechanism of the immobilized GOD toward glucose. Figure
Some representative experimental data from graphene based biosensors. (a) Graphene based glucose biosensor: (i) O2 saturated PBS solution without glucose and (ii) O2 saturated PBS solution with different concentrations of glucose. (b) Graphene based cholesterol biosensor: (i) 0.25
Cholesterol and its esters are the essential components found in the cell membranes of all human and animal cells. The normal cholesterol limit in human serum is in the range of 1.0–2.2 mM and its excessive accumulation in blood results in fatal diseases. Gholivand and Khodadadian [
A novel H2O2 biosensor was fabricated using graphene/Fe3O4-AuNP and graphene/Fe3O4-AuNP nanocomposites coated with horseradish peroxidase [
The sensitivity, selectivity, and detection limit of the enzymatic electrode are impressive. However, these electrodes suffer from the limitation of reproducibility, high cost, and complexity in enzyme immobilization procedure. The enzymatic electrodes are also very sensitive towards the change in pH of the solution, temperature, and toxic chemicals. In order to overcome these limitations, the use of nonenzymatic biosensors in the detection of biomolecules has been introduced as discussed below.
Nonenzymatic detection of biomolecules using graphene-based electrodes has attracted significant attention due to its low fabrication cost, high sensitivity, and long-term stability. Li et al. [
DNA biosensor was fabricated using graphene/PANI nanocomposite films [
In the development of tissue engineered constructs, the need for reliable and sensitive tools to assess the artificial tissue environment has become vitally important. Such platforms require to be constantly monitored in terms of various physiologically relevant parameters to evaluate the functionality of the engineered tissue constructs. Microfluidic systems are able to mimic various signals that direct cell fate to create specific organ constructs by precise control of the chemical and mechanical stimuli at microscale [
Recently, a number of studies have been reported on the combination of biosensor capabilities in microfluidic devices for tissue engineering. Weltin et al. presented a multiparametric microphysiometry platform to monitor the metabolism of T98G human brain cancer cells cultured in dynamic flow conditions [
A rapid translation of successful biosensing technologies to tissue engineering platforms is only at an early stage due to several challenges that researchers have to face for a proper integration in microfluidic systems [
On one hand, POC systems often present several capabilities in a single device, such as fluid handling, sample preparation (concentration, washing, etc.), and the possibility to perform different reactions for biochemical assays. On the other hand, the miniaturization and integration in microfluidic cell culture systems of these capabilities can be very challenging from a fabrication and operational point of view, although several attempts are reported [
There has been a growing interest in biosensor research for applications in tissue engineering. However, the progress has remained limited. Even though numerous optical, electrochemical, magnetic, acoustic, thermometric, and piezoelectric sensors have been reported in the literature and, often, are already available in the market, showing great sensitivity and sensibility, the most successful among them in tissue engineering applications have been the electrochemical and optical ones, while the thermometric and magnetic transductions have failed to have any practical impact. The challenges for widespread applications of biosensors in tissue engineering include their miniaturization and integration in microfluidic systems. The continuous real time monitoring of analytes in tissue engineering is still at an early stage and can bring enormous possibilities in the field. The creation of microfluidic tissue engineering platforms with automated, sensitive, and real-time monitoring capabilities will hugely benefit the translation of such systems to clinics, as the full assessment of their parameters is a must for clinical applications. The successful widespread application of biosensors in tissue engineering particularly on microfluidic platforms will require standardization of the systems and the processes.
The authors declare that there is no conflict of interests regarding the publication of this paper.