BIOSENSORS
Biosensor is an analytical device containing an immobilized biological material (enzyme, antibody, nucleic acid, hormone, or whole cell) that
can specifically interact with an analyte and produce physical, chemical, or electrical signals that can be measured.
Professor Leland C Clark is the father of biosensors.
Biosensors involve quantitatively analyzing the concentration of various substances or chemicals by converting their biological actions into
measurable signals.
The term “biosensor” is short for “biological sensor. The word biosensor indicates the combination of two parts: Bio element and Sensor
element. Biosensors are devices that combine a biological component to
detect an analyte and a physicochemical component to produce a signal
which is measurable.
Components of biosensors:
a) Biological component (enzymes, cells, etc.)
b) Physical component (transducer, amplifier)
WORKING OF BIOSENSORS
ANALYTE: an analyte is a compound (e.g., glucose, urea, drug, pesticide)
whose concentration has to be measured.
BIOMATERIAL: It is an element in which the analyte is bound. (e.g., tissues,
cells, organelles, etc)
TRANSDUCER: It is a device that converts energy from one form to another
form.
AMPLIFIER: which enhances the signal.
PROCESSOR: analyses the signal and forms the output
DISPLAY: It shows the output.
PRINCIPLE OF BIOSENSORS
Biosensors operate on the principle of integrating a biological recognition element with a transducer to detect and quantify specific analytes. The biological
sensing element interacts selectively with the target analyte, inducing a change in its physicochemical properties. This change is transduced by the transducer
into a measurable signal, such as electrical current, voltage, light intensity, or frequency. The signal is then amplified and detected, allowing for the
quantification of the analyte. Biosensors find wide-ranging applications in pharmaceutical biotechnology, including drug discovery, pharmacokinetics, and point-
of-care diagnostics, due to their ability to provide rapid, sensitive, and selective detection of biomolecules in various biological samples. Calibration and
validation are essential steps in ensuring the accuracy, precision, and reliability of biosensor measurements. Overall, biosensors play a crucial role in advancing
biomedical research and improving patient care outcomes by facilitating real-time monitoring and analysis of biological processes.
A GOOD BIOSENSORS
A good biosensor should possess the following characteristics to ensure high performance and reliability:
1. Selectivity: The ability to specifically recognize and bind to a particular analyte, even in the presence of other substances.
2. Reproducibility: The ability to generate identical responses for similar experimental setups, ensuring precision and accuracy.
3. Stability: Resistance to ambient disturbances, maintaining a consistent response over time.
4. Sensitivity: The ability to detect low concentrations of analytes, with a high limit of detection.
5. Linearity: The ability to produce a response that is proportional to the concentration of the analyte, ensuring accurate quantification.
Additionally, biosensors should be:
Cost-effective
Free from electrical noise.
Cheap, small, portable, and capable of being used by semi-skilled operators
The reaction should be independent of physical parameters (stirring, pH, and temperature)
Rapid and continuous in measurement
Require minimal reagents for calibration
Have a fast response time
Be able to measure non-polar molecules
Be biocompatible and reliable
Have a small size
Be easy to use and maintain
Be able to detect dangerous biological agents or chemicals within the human body.
TYPES OF BIOSENSORS
1. Electrochemical Biosensor: a) Amperometric Biosensor
b) Potentiometric Biosensor
c) Conductometric Biosensor
2. Thermometric Biosensor
3. Optical Biosensor: a) Fiber Optic Lactate Biosensor
b) Optical Biosensors for Blood Glucose
4. Piezoelectric Biosensors
5. Whole-cell Biosensor
6. Immuno-Biosensor
Electrochemical Biosensor
An electrochemical Biosensor is a simple device. It measures the measurement of electronic current, ionic or by conductance changes carried by bioelectrodes.
Many chemical reactions produce or consume ions or electrons which in turn cause some change in the electrical properties of the solution which can be sensed
out and used as measuring parameter.
This operates at fixed potential with respect to references electrode involved the detection of the current generated by the oxidation or reduction of species at the
surface of the electrode. Electrochemical biosensor is mainly exploited for the recognition of hybridized DNA, glucose concentration, DNA binding drugs. On the
basis of measuring electrical parameters electrochemical biosensor can be classified as A) Amperometric B) Conductometric C) Potentiometric.
Electrochemical biosensors offer several advantages, including high sensitivity, rapid response, and the ability to perform real-time measurements.
Amperometric biosensors
Amperometric biosensors operate based on the movement of electrons resulting from enzyme-catalyzed redox reactions, providing a means for detecting and
quantifying specific analytes. These biosensors typically employ enzyme-electrode or chemically modified electrodes to facilitate the redox reactions.
In the enzymatic reaction, the substrate undergoes oxidation or reduction, with the electrons being transferred directly to the electrode surface via the enzyme
and a redox mediator. This transfer of electrons results in a measurable electric current, the magnitude of which is proportional to the concentration of the
substrate. One of the most common Amperometric biosensors is the glucose biosensor.
In the first generation Amperometric biosensors (described above), there is a direct transfer of the electrons released to the electrode which may pose some
practical difficulties. A second generation Amperometric biosensors have been developed wherein a mediator (e.g. ferrocenes) takes up the electrons and then
transfers them to electrode. These biosensors however, are yet to become popular.
The working principle of a blood-glucose biosensor involves the use of a glucose oxidase enzyme to catalyze the oxidation of glucose, resulting in the production
of hydrogen peroxide (H2O2). This reaction is coupled with the reduction of oxygen at the electrode surface, generating an electrical current that is proportional
to the glucose concentration in the blood sample.
Here's a step-by-step explanation of how a blood-glucose biosensor works:
1.Enzyme Immobilization: Glucose oxidase enzyme is immobilized onto the surface of a working electrode, typically made of a conductive material such as
platinum or carbon. The enzyme is selectively specific to glucose and catalyzes its oxidation reaction.
2.Glucose Oxidation: When a blood sample containing glucose is applied to the biosensor, the glucose molecules diffuse to the surface of the working electrode.
The glucose oxidase enzyme catalyzes the oxidation of glucose, converting it into gluconic acid and producing hydrogen peroxide (H2O2) as a byproduct.
3.Electrochemical Reaction: The generated hydrogen peroxide
undergoes an electrochemical reaction at the electrode surface.
Specifically, it is oxidized at the electrode, resulting in the transfer of
electrons and the reduction of oxygen present in the surrounding
solution.
4.Current Generation: The electrochemical reaction involving hydrogen
peroxide and oxygen generates an electrical current, which is
measured by the biosensor. The magnitude of the current is directly
proportional to the concentration of glucose in the blood sample.
5.Data Display: The measured current is converted into glucose
concentration units and displayed on a digital screen or readout,
providing the user with real-time information about their blood
glucose levels.
Major drawbacks of first generation glucose biosensors:
Amperometric measurement of hydrogen peroxide required a high operating potential (0.6 V) for high selectivity.
Restricted solubility of oxygen in biological fluids, which produced fluctuations in the oxygen tension.
Deactivation of the enzyme due to the production of hydrogen peroxide.
Potentiometric biosensors
Unlike Amperometric sensors, which measure current, potentiometric sensors measure the electrical potential (voltage) between a working electrode and a
reference electrode. This potential difference arises due to the ionic activity of the solution, influenced by specific interactions between the target molecule and
the biorecognition element. pH Biosensors, ion-selective field effect transistors(ISFET), gas biosensors are good examples.
Working:
1.Electrode Configuration: Potentiometric biosensors consist of two main electrodes: a working electrode and a reference electrode. The working electrode is
typically coated or modified with a recognition element, such as enzymes, antibodies, or ion-selective membranes, which selectively interact with the analyte of
interest. The reference electrode provides a stable reference potential against which the potential at the working electrode is measured.
2.Ion Selective Membrane: In some potentiometric biosensors, an ion-selective membrane is used to enhance selectivity for specific ions or analytes. The ion-
selective membrane allows only certain ions to pass through, resulting in a potential difference across the membrane proportional to the concentration of the
target analyte.
3.Ion Exchange: When the analyte of interest interacts with the recognition element or ion-selective membrane, it causes a change in the distribution of ions
near the working electrode surface. This change in ion concentration leads to a change in electrical potential or voltage at the working electrode.
4.Potential Measurement: The potential or voltage difference between the
working electrode and the reference electrode is measured using a voltmeter or
potentiostat. This measured potential is then correlated with the concentration
of the analyte in the sample.
5.Calibration: Before use, potentiometric biosensors are typically calibrated using
standard solutions with known analyte concentrations. Calibration curves are
constructed to establish the relationship between the measured potential and
the analyte concentration, allowing for accurate quantification of the analyte in
unknown samples.
Conductimetric biosensors
There are several reactions in the biological systems that bring about changes in the ionic species. These ionic species alter the electrical conductivity which can
be measured. Conductometric biosensors are a type of biosensor that detect changes in the electrical conductivity of a solution resulting from biochemical
reactions or interactions between the analyte and the biological recognition element.
Working Principle:
Conductometric biosensors measure changes in the electrical conductivity of a solution due to the presence of ions produced or consumed during a biochemical
reaction. When the target analyte interacts with the biological recognition element (e.g., enzyme, antibody, or DNA), it leads to changes in the ion concentration
or solution conductivity. These changes are then detected and quantified by measuring the electrical resistance or conductance of the solution.
A good example of conductimetric biosensor is the urea biosensor utilizing immobilized urease. Urease catalyses the following reaction.
The above reaction is associated with drastic alteration in ionic concentration which can be used for monitoring urea concentration. In fact, urea biosensors
are very successfully used during dialysis and renal surgery.
Thermometric Biosensor
There are many more biological reactions connected with the production of heat and it forms the basis of
thermometric Biosensors.
Thermometric biosensors measure changes in temperature that occur during a biochemical reaction
catalyzed by the biological recognition element. When the target analyte interacts with the recognition
element (e.g., enzyme, antibody, or nucleic acid), it initiates a reaction that either generates or consumes
heat. This leads to a change in temperature, which is then detected and quantified using a temperature
sensor. The difference in the temperature between the substrate and product is measured by thermistors
(thermal biosensors).
Calorimetric microsensors have been used for detection of cholesterol in blood serum based on
production of heat by enzymatic reaction.
They are more commonly referred to as thermal biosensors or calorimetric biosensors.
Example: a) biosensors for detection of serum cholesterol.
b) biosensors for estimation of urea, uric acid, penicillin G
These biosensors can be used as a part of ELISA and the new technique is referred to as thermometric
ELISA (TELISA).
A diagrammatic representation of a thermal biosensor is depicted in Fig. beside. It consists of a heat
insulated box fitted with heat exchanger (aluminum cylinder). The reaction takes place in a small enzyme
packed bed reactor. As the substrate enters the bed, it gets converted to a product and heat is generated.
The difference in the temperature between the substrate and product is measured by thermistors. Even a
small change in temperature can be detected by thermal sensors.
Optical Biosensor
The optical Biosensors are safe for non-electrical remote sensing of materials.
Another advantage is the Biosensors do not require any reference sensors as the comparative signals are generated by using the same source of light as the
sampling sensor.
Optical biosensors are the devices that utilize the principle of optical measurements (absorbance, fluorescence, chemiluminescence etc.). They employ the use
of fibre optics and optoelectronic transducers. The word optrode, representing a condensation of the words optical and electrode, is commonly used. Optical
biosensors primarily involve enzymes and antibodies as the transducing elements.
Optical biosensors allow a safe non-electrical remote sensing of materials. Another advantage is that these biosensors usually do not require reference sensors,
as the comparative signal can be generated using the same source of light as the sampling sensor. Some of the important optical biosensors are briefly
described hereunder.
Principles of Operation:
Optical biosensors function based on the interaction between light and a biological recognition element, such as enzymes, antibodies, or nucleic acids. These
interactions induce measurable changes in the optical properties of the sensor, which are then correlated with the concentration of the target analyte.
Key Components:
1. Biological Recognition Element: This element selectively binds to the target analyte, initiating the sensor's response.
2. Transducer: Converts the biochemical signal generated by the recognition event into an optical signal. Common transducers include surface plasmon
resonance (SPR), fluorescence, and photonic crystals.
3. Optical Detection System: Detects and quantifies changes in the optical signal, providing information about the presence and concentration of the analyte.
• The principle of piezoelectric biosensors is used in sound vibrations, hence it is called acoustic Biosensors.
• The crystals with positive and negative charges vibrate with characteristic frequencies. Adsorption of certain molecules on the crystal
surface alters the resonance frequencies which can be measured by electronic devices.
• Examples: A piezoelectric biosensor for organophosphorus insecticide.
A biosensor for cocaine in gas phase.
Principles of Operation:
Piezoelectric biosensors typically consist of a piezoelectric material, such as quartz or a piezoelectric polymer, coated with a biological
recognition element, such as antibodies, enzymes, or nucleic acids. When the target analyte binds to the recognition element on the sensor
surface, it induces a change in the mass, density, or viscoelastic properties of the sensor layer. This change results in a proportional alteration
in the resonant frequency or amplitude of the piezoelectric device, which can be measured and correlated with the concentration of the
analyte.
Key Components:
1. Piezoelectric Material: The core component of the sensor, capable of converting mechanical stress into an electrical signal and vice
versa.
2. Biological Recognition Element: Selectively binds to the target analyte, initiating the mass or viscoelastic changes on the sensor surface.
3. Transducer and Electronics: Convert the mechanical response of the piezoelectric material into an electrical signal for detection and
analysis.
