Biomedical Instrumentation: Ms. Divya B AP/BME

Biomedical Instrumentation
Ms. Divya B AP/BME

• A biosensor is an analytical device which is used to
determine the presence and concentration of a specific
substance in a biological analyte
2 v 1.2
3 v 1.2
Amperometric biosenor
• Amperometric biosensors are self-contained integrated devices
based on the measurement of the current resulting from the
oxidation or reduction of an electroactive biological element
providing specific quantitative analytical information
4 v 1.2
Amperometric Glucose Biosensor
 Developed by Updike and Hicks
 Enzyme Glucose oxidase catalyze the oxidation of glucose by
• molecular oxygen producing glucolactone and hydrogen
peroxide.
 In order to work as a catalyst, GOx requires a redox cofactor –
flavin adenine dinucleotide (FAD), works as an initial electron
acceptor and is reduced to FADH2.
•Glucose + GOx –FAD+ Glucolactone+ GOx – FADH2
5 v 1.2
• The cofactor is regenerated by reacting with oxygen, leading to
the formation of hydrogen peroxide
• GOx – FADH2 + O2 GOx – FAD +
H2 O 2
• Hydrogen peroxide is oxidized at a platinum electrode. The number of

electron transfers, at electrode surface is directly proportional to the number
of glucose molecules present in the blood.
• H 2 O2 2H+ +O2 + 2 e-
• Three strategies used for the electrochemical sensing of glucose are

 By measuring oxygen consumption
 By measuring the amount of hydrogen peroxide produced by the
enzyme reaction
 By using a diffusible or immobilized mediator to transfer the electrons
from Gox to the electrode.
6 v 1.2
7 v 1.2
Generations
 1st generation: the normal product of the reaction
diffuses to the transducer and causes electrical
response
 2nd generation: involves specific mediators between
reaction and transducer to generate improved response
 3rd generation: reaction itself causes the response
8 v 1.2
First generation glucose
biosensors
The first generation glucose biosensors estimated glucose concentration in the
sample based on hydrogen peroxide production by glucose oxidase utlizing
dissolved oxygen.
• Based on this technology, Yellow spring Instrument company, launched the

first commercial glucose biosensor in market in 1975 for the direct
measurement of glucose.
• The usage of the most expensive metal platinum for the fabrication of
9 this v 1.2
electrode restricted the biosensor to clinical laboratories only.

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.
10 v 1.2
Second generation glucose
•
biosensor
The second generation glucose biosensor utilized redox mediator to transfer
electrons from the enzyme to the working electrode surface.
• A variety of redox mediators, such as ferrocene, ferricyanide, quinines, methylene

blue etc were used to improve sensor performance.
• Usage of redox mediator eliminated the need of oxygen for electron transfer at the
electrode surface, thus overcoming the drawback of limited oxygen pressure observed
in the first generation biosensor.
• The lower redox potential of chosen mediators (0-2 V) results in no interference from
other electroactive species such as uric acid, ascorbic acid.
• Redox mediator enhances the electron

11 transfer between the redox center of enzyme
v 1.2
and the electrode surface.
Major drawbacks of second generation
glucose biosensors
• High competition between redox mediator and oxygen.
• Interference of other electroactive species lead to false

and inaccurate results.
• Small size and highly diffusive nature of mediators poses

problem of leaching of mediator from intermediate
region between enzyme and electrode surface.
12 v 1.2
Third generation glucose
biosensors
• The third generation glucose

biosensors are based on the direct
electron
• transfer between the active center of enzyme and
the electrode.
• The intrinsic barrier to electron flow is the globular

buried active site of
structure
enzyme.of glucose oxidase with the active site,
containing FAD/FADH2 redox cofactor, buried deep inside a
cavity of ~ 13 A◦ is a major hinderance for direct electron transfer.
13
• Carbon nanotubes immobilized electrode surface
v 1.2
provide suitable orientation for enzyme

Non-enzymatic glucose
biosensors
• The use of non-enzymatic electrodes as glucose sensors potentially promises a fourth
generation to analytical glucose oxidation.
• The active metal nanoparticle undergo a oxidation step that forms a hydrous
14 oxide layer OHads that mediate oxidation
v 1.2
of the adsorbed species.
Glucose Biosensors Based on
Carbon Nanotube Nano electrode
Ensembles
 The development of glucose biosensors based on carbon nanotube (CNT)
Nano electrode ensembles (NEEs) for the selective detection of glucose.
 CNTs have a high electrocatalytic effect and a fast electron-transfer rate.
 Glucose oxidase was covalently immobilized on CNT NEEs via
carbodiimide chemistry by forming amide linkages between their amine
residues and carboxylic acid groups on the CNT tips.
 The catalytic reduction of hydrogen peroxide liberated from the enzymatic
reaction of glucose oxidase upon the glucose and oxygen on CNT NEEs
leads to the selective detection of glucose.
 The biosensor effectively performs a selective electrochemical analysis of
glucose in the presence of common interferents (e.g., acetaminophen, uric
and ascorbic acids), avoiding the generation of an overlapping signal from
such interferers. Such an operation eliminates the need for permselective
membrane barriers or artificial electron mediators, thus greatly simplifying
the sensor design and fabrication.
15 v 1.2
• The fabrication of glucose biosensors based on CNT
for the and sensitive detection NEEs
selective eliminate potential interference throughof the glucose.
preferential
CNT
NEEs
detection of hydrogen peroxide. Such development of
interference-free transducers should simplify the design and
fabrication conventional and miniaturized sensing probes.
The
of glucose biosensor based on an aligned CNT NEE is thus
suitable for the highly of selective detection of glucose in a sweat,
variety Thebiological fluids (e.g., urine, and
serum). biosensor fabrication saliva,
technology demonstrated
in work is readily applicable to fabrication of
this
biosensors based on oxidases, the as biosensors other
cholesterol, alcohol, lactate, acetylcholine,
such choline, for
anthine, and xanthine. hypox-
16
Glucose biosensor test
strips
Dry coating of GO +
Fc
e’s electrodes
Meter
Read
glucose
Patient adds drop of blood,
then inserts slide into
meter
Patient reads glucose level on
I meter
17 v 1.2
Thank you
18 v 1.2

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Biomedical Instrumentation

Ms. Divya B AP/BME

•Glucose + GOx –FAD+ Glucolactone+ GOx – FADH2

• Hydrogen peroxide is oxidized at a platinum electrode. The number of

• Three strategies used for the electrochemical sensing of glucose are

• Based on this technology, Yellow spring Instrument company, launched the

electrode restricted the biosensor to clinical laboratories only.

• Amperometric measurement of hydrogen peroxide

• Restricted solubility of oxygen in biological fluids,

• Deactivation of the enzyme due to the production of hydrogen

• A variety of redox mediators, such as ferrocene, ferricyanide, quinines, methylene

• Redox mediator enhances the electron

• High competition between redox mediator and oxygen.

• Interference of other electroactive species lead to false

• Small size and highly diffusive nature of mediators poses

• The third generation glucose

• The intrinsic barrier to electron flow is the globular

provide suitable orientation for enzyme

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