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BIOSENSORS
CONTENTS
Introduction
Basic components of Biosensor
Working of Biosensor
Types of Biosensor
Applications of Biosensor
Conclusion
DEFFINITION
A device that uses specific biochemical reactions mediated
by isolated enzymes, immunosystems, tissues, organelles or
whole
cells
to
detect
chemical
compounds
usually
by
electrical, thermal or optical signals.
0r
The term biosensor is defined as a sensor incorporating
biological elements such as enzymes, antibodies, receptors
proteins, nucleic acids, cells, or tissue sections - as the
recognition element, coupled to a transducer.
Block Diagram of a Biosensor
Sample
(Analyte or
Substrate)
Biorecognition
Element
Transducer
Signal
Processing
Device
PRINCIPLE
• The biological material is immobilized and a contact is made
between the immobilized biological material and the
transducer
• The analyte binds to the biological material to form a bound
analyte which in turn produces the electronic response that
can be measured.
• Sometimes the analyte is converted to a product which could
be associated with the release of heat, gas (oxygen),
electrons or hydrogen ions. The transducer then converts the
product linked changes into electrical signals which can be
amplified and measured
WORKING PRINCIPLE
• Biosensors basically involve the quantitative analysis of
various substances by converting their biological actions
into measurable signals.
• Generally the performance of the biosensors is mostly
dependent on the specificity and sensitivity of the
biological reaction, besides the stability of the enzyme.
WORKING PRINCIPLE
Analyte diffuses from the solution to the surface of the
Biosensor.
Analyte reacts specifically & efficiently with the Biological
Component of the Biosensor.
This reaction changes the physicochemical properties of
the Transducer surface.
This leads to a change in the optical/electronic properties of
the Transducer Surface.
The change in the optical/electronic properties is
measured/converted into electrical signal, which is detected.
Analyte
Response
Detection
Signal
Analysis
Sample
handling/preparation
9
10
11
12
(e)
(f)
(d)
(a)
(b)
(c)
The interaction of the analyte (a) with the bioreceptor, which identifies
the stimulus (b) is designed to produce an effect measured by the
transducer (c), which converts it to an electrical signal. The output from
the transducer is amplified (d), processed (e) and displayed (f).
IDEAL BIOSENSORS
The output signal must be relevant to measurement
environment.
The functional surface must be compatible with the
transducer.
High specificity and selectivity (low interference).
Sufficient sensitivity and resolution .
Sufficient accuracy and repeatability
Sufficient speed of response
Sufficient dynamic range.
Insensitivity to environmental interference or their
effects must be compensated
Biosensor components
1. Analyte
2. Biological elements
3. Transducer
4. (processor)
5. (monitor)
ANALYTE
THE ANALYTE
(What do you want to detect?)
Molecule
Protein, toxin, peptide, vitamin, sugar, metal ion
Cholera toxin
Glucose
BIOLOGICAL ELEMENTS
BIOLOGICAL ELEMENTS
Fab
Active site
Membrane receptors
Competitive binding
Fc
Antibody
Enzyme
Cell
Polymer/Hydrogel
Biological recognition element• It is the sensitive biological element or biological material (tissue
,microorganisms, organelles,cell receptors , enzymes, antibodies,
nucleic acids,etc.) or bio- mimetic component
that
interacts
(binds or recognises) the analyte under study.
•
The biologically sensitive elements can also be
created by biological engineering.
Biological elements
1. Enzymes
2. Antibodies
3. Hormone receptors
4. Cells
5. Cell organelle
6. Tissues
7. Membranes
8. Micro organisms
9. Nucleic acids
10. Biomimetic materials
1.EnzymesEnzyme-based biosensors use their catalytic
activity and binding capabilities for specific
detection .
The catalytic activity of the enzymes
provides these types of biosensors with
the ability to detect much lower limits
than with normal binding techniques.
This catalytic activity is related to the
integrity of the native protein structure.
Most common – Glucose oxidase, Urease, Alcohol oxidase etc.
Commercial example is GLUCOSE SENSOR using oxidase (GOD) .
Clarks (1962)
Glucose + O2 -- glusoce oxidase  Gluconic acid + H2O2 (1)
H2O2  O2 + 2H+ + 2eO2 + 4H+ + 4 e-  2H2O
-and current flows.
(3) at anode
(2) at cathode
There are three measurement routes-pH change (acid production)
- O2 consumption (fluorophore monitor)
- H2O2 production (electrochemical)
2. Micro-organisms –
Microorganisms such as bacteria and fungi
can be used as biosensors to detect specific
molecules or the overall state of the
surrounding environment.
Cell behaviour such as cell metabolism,
cell viability, cell respiration, and bioluminescence can be used as indicators
for the detection.
Furthermore, proteins that are present in
cells can also be used as bio-receptors for the
detection of specific analytes.
Rhodococcus erythropolis (in collagen) used
to measure BOD.
3. Cell / Tissue samples.
Use of cell as a biosensor occurred in 1977 by
Rechnitz .
Rechnitz coupled Streptococcus faecium on the
surface of an ammonia gas sensing membrane
electrode .
This Rechnitz electrode was capable of detecting
the amino acid ARGININE.
4. Organelles – Mitochondra , Cell walls etc.
5. Membranes .
6. Bio-mimetic materials A bio mimetic biosensor is an artificial or
synthetic sensor that mimics the function of a
natural biosensor .
These can include aptasensors , where apta
-sensors use aptamers as the biocomponent.
Aptamers are synthetic single
stranded
nucleic acid that can be designed to
identify
or
recognize
amino-acids,
oligosaccharides , peptides , and proteins .
Aptamers have high affinity , high selectivity,
cheaper & easy to synthesize .
B. Affinity biosensors:
device in which receptor molecules bind analyte molecules
causing physicochemical change that is detected by a transducer.
Receptors used are-
1. Antibody / Antigen (Immunosensor) The high specificity between an antibody and
an antigen can be utilized in this type of sensor
technology. Biosensors utilizing this specificity
must ensure that binding occurs under conditions
where nonspecific interactions are minimized .
Binding can be detected either through
fluorescent labelling or by observing a
refractive index or reflectivity changes.
2. Hormone receptor/Antagonist-
3. Nucleic acid -The complementary relationships between nucleic acid
‘bases’ in the DNA form the basis of specificity in
nucleic acid based biosensors.
These sensors are capable of detecting trace amounts of
microorganism DNA by comparing it to a complementary
strand of known DNA.
By unwinding the target DNA strand, adding the DNA
probe, and annealing the two strands , the probe
will hydrolyze to the complementary sequence on the
adjacent strand . If the probe is tagged with a
fluorescent compound , then this annealing can be
visualized under a microscope.
eg. DNA Chip.
(Target probe )
(Capture probe)
General DNA biosensor scheme--Target DNA is captured at the recognition layer (A),
and the resulting hybridization is transduced into a measurable electronic signal (B).
It is used for genome mutation detection & clinical diagnosis.
BIOLOGICAL ELEMENTS CONSTRUCTION
 The principle of detection is the specific
binding of the analyte of interest to the
complementary biorecognition element
immobilised on a suitable support medium.
BIOLOGICAL ELEMENTS CONSTRUCTION
I.
Recognition element immobilization (Immobilization of biological
receptor)Biological receptors, i.e. enzymes, antibodies, cells or tissues with high
biological activity, can be immobilized in a thin layer at the transducer
surface by using different procedures.
(a) Entrapment behind a membrane - viscous aqueous solution
trapped by membrane permeable to analyte.
(b) Entrapment of biological receptors within a polymeric matrixMembranes—
Cellophane, Cellulose acetate, Polyurethane membranes .
Gel entrapment–
Agarose, Gelatin, Agar gel , Polyacrylamide gel.
MicroencapsulationEncapsulation inside Liposomes, or absorbed in fine carbon
particles that are incorporated in a gel or membrane
.
(c) Entrapment of biological receptors within
self-assembled monolayers (SAMs)
or bilayer lipid membranes (BLMs).
(d) Adsorption:
direct adsorption onto membrane or transducer;
can also be adsorbed onto pre -adsorbed proteins .
(e) Covalent binding (via –COOH, -NH2, -OH ) , or cross
linking (via glutaraldehyde ) to transducer or
membrane surface.
Receptors are immobilized either alone or are mixed with other
proteins, such as bovine serum albumin (BSA), either directly on
the transducer surface, or on a polymer membrane covering it .
SIGNAL
(How do you know there was a detection?)
Specific recognition?
PRINCIPLE OF DETECTION
 The principle of detection is the specific binding of the analyte of
interest to the complementary biorecognition element
immobilised on a suitable support medium.
 The specific interaction results in a change in one or more physicochemical properties (pH change, electron transfer, mass change,
heat transfer, uptake or release of gases or specific ions) which are
detected and may be measured by the transducer.
TRANSDUCER-
The transducer or the detector element
transforms the signal resulting from the
interaction of the analyte with the biological
recognition element into another signal that
can be more easily measured and quantified.
AMPLIFIER, MICROPROCESSOR AND DISPLAY-
Biosensor reader device with the associated
electronics or signal processors that are
primarily responsible for the display of the
results.
TYPES OF BIOSENSOR
• Biosensor is broadly classified into two classes:
• I) On the basis of biological element
• a) Enzyme Biosensor
• b) Microbial Biosensor
• c) Antibody Based.
II) ON THE BASIS OF TRANSDUCING
ELEMENT
 Calorimetric/Thermal Detection Biosensors.
 Optical Biosensors.
 Resonant Biosensors.
 Piezoelectric Biosensors.
 Ion Sensitive Biosensors.
 Electrochemical Biosensors.
 Conductimetric Sensors.
 Amperometric Sensors.
 Potentiometric Sensors.
Calorimetric / Thermal Detection Biosensors.
Uses Absorption / Production of Heat.
Total heat produced/absorbed is ᾶ Molar Enthalpy/Total
No. of molecules in the rn.
Temp. measured by Enzyme Thermistors.
Advantages:
• No need of Frequent recalibration.
• Insensitive to the Optical & Electrochemical Properties of
the sample.
Uses:
Detection of: (1) Pesticides .
(2) Pathogenic Bacteria.
Optical Biosensors.
Colorimetric for colour - Measures change in Light
Adsorption.
 Photometric for Light Intensity - Detects the Photon
output.
Resonant Biosensors.
An Acoustic Wave Transducer is coupled with Bioelement.
Measures the change in Resonant Frequency.
Piezoelectric Biosensors.
Uses Gold - To detect specific angle at which ȇ waves are
emitted when the substance is exposed to laser
light/crystals like quartz, which vibrates under the influence
of an electric field.
 Change in Frequency ᾶ Mass of Absorbed material.
Ion Sensitive Biosensors.
Are semiconductor with ion-sensitive surface.
Surface Electrical Potential changes when the ions &
semiconductors interact.
Measures the Change in Potential.
Uses:
opH Detection.
Electrochemical Biosensors.
Underlying Principle – Many chem.rns produce or consume
ions or ȇs causing some change in the elctrical properties of
the solution that can be sensed out & used as a measuring
parameter.
Uses:
Detection of :
oHybridized DNA
oDNA- binding Drugs &
oGlucose Concentration.
Conductimetric Sensors.
Measures Electrical Conductance/Resistance of the
solution.
Conductance Measurements have relatively Low Sensitivity.
 Amperometric Biosensors.
 High Sensitivity Biosensor.
 Detects electroactive species present in the biological
test samples.
 Measured Parameter – Current.
Potentiometric Sensors.
Working Principle – When ramp voltage is applied to an
electrode in solution, a current flow occurs because of
electrochemical reactions.
Measured Parameter – Oxidation / reduction Potential of
an Electrochemical rn.
APPLICATIONS
1. MEDICAL
a. Biosensors are used in both clinical and laboratory use in medical
care.
Glucose monitoring in diabetes patients .
Medtronic glucose sensor - implants in major vein of heart.
b. Tumor cells are used as biosensors to monitor the susceptibility of
chemotherapeutic drugs.
c. Routine analytical measurement of folic acid, biotin, vitamin B12 and
pentothenic acid .
d. Micro- and nanoscale biosensors—
Genome mutation detection , cancer detection & clinical
diagnosis.
Bacterial-UTI , Human Immunodeficiency Virus (HIV)
Detection,
Hepatitis and Anthrax detection.
2. BIO / PHARMACEUTICAL
RESEARCH
a. Quality assurance
b. Study of biomolecules and their interaction
c. Protein engineering .
d. Drug discovery , evaluation monitor the manufacturing
of biological activity of new compounds (research field).
e. Biosensors are used for measuring concentration of various metal
ions by specific protein concentration or by using genetically
modified organisms.
f. Aptamers are used for the detection of proteins –Thrombin , IgE ,
HIV – tat protein , Lysozyme , toxin.
2. BIO / PHARMACEUTICAL RESEARCH
g. Biosensors are used in monitoring of the glutamate and acetyl
choline , which is the main cause in neurodegenerative diseases.
h. Microbiology: bacterial and viral analysis .
i. Biosensors are used in analysing micro dialysis samples.
j. Biosensors are used in biotechnological process such as to
determine proteins or peptides.
k. Biosensors are also used in determining intracellular proteins and
also plasmids.
l. Detection of cancer biomarkers – CEA , PSA , CA-125 &
Tumour necrosis factor .
3. INDUSTRIAL / AGRICULTURAL
a. Biosensors used in process control will be able to
measure materials present in the process.
b. Use of biosensors in industry will improve
manufacturing techniques, this will allow for usage of
wider variety of sensing molecules.
c. Biosensors are used in controlling the industrial
processes.
d. Microbial sensor measure Ammonia & Methane.
4. ENVIRONMENTAL
a. Biosensors are used in detecting environmental pollutants
and monitoring of Mines, Industries and toxic gases.
b. Biosensors are used in the BOD measurement during waste
water treatment.
c. Biosensors are used in the detection of poly aromatic
hydrocarbons present in water.
d. Environmental applications e.g. the detection of pesticides
and river water contaminants such as heavy metal ions.
e. Detection and determination of organophosphates .
5. FOOD INDUSTRY
a. Quality assurance in food industries , ex. E. Coli, Salmonella.
b. Food & drink production analysis.
c. Biosensors are used for detection of food freshness marker
determining parameters in wine industry.
d. Determination of drug residues in food, such as antibiotics
and growth promoters, particularly meat and honey.
e. Detection of toxic metabolites such as mycotoxins.
Example of biosensors
Pregnancy test - Detects the hCG(human chorionic
gonadotropin) protein in urine.
Glucose monitoring device (for diabetes patients)
Monitors the glucose level in the blood.
Infectous disease biosensor from
RBS
Old time coal miners’ biosensor
6. BIODEFENCE
a. Detection of pathogens.
b. Remote sensing of airborne bacteria / virus ,
e.g. In counter bio-terrorist activities.
c. Detection system for biological welfare agent
eg. Bacillus anthracis (anthrax) spores.
d. Determining levels of toxic substances before and
after bioremediation.
e. Crime detection.
GLUCOSE OXIDASE SENSOR
• A common example of a commercial biosensor is the blood
glucose biosensor, which uses the enzyme glucose oxidase to break
blood glucose down.
• In doing so it first oxidizes glucose and uses two electrons to reduce the
FAD (a component of the enzyme) to FADH2.
• This in turn is oxidized by the electrode (accepting two electrons from
the electrode) in a number of steps.
• The resulting current is a measure of the concentration of glucose. In
this case, the electrode is the transducer and the enzyme is the
biologically active component.