EEE 529 Microsystems Amperometric Biosensors

Agamyrat Agambayev 520112003

Contents:1. Introduction to Biosensors2. Definitions3. Amperometric Biosensors4. Generations of Amperometric Biosensors5. Performance factors6. Applications7. References

MULTIDISCIPLINARY NATURE OF BIOSENSOR:

INTRODUCTIONBIOSENSORS :Compact analytical devices that bring together the use of a biological, a biologically-derived or a biomimic element to recognize the analyte.

INTRODUCTIONBiosensor = biorecognation element + transducer. 

Biorecognation elementa biomolecule that recognizes the target analyte Transducer converts the recognition event into a measurable signal

The uniqueness of a biosensor is that the two components areintegrated into one single sensor 

INTRODUCTIONBiosensor = biorecognation element + transducer. 

Biosensor = biorecognation element + transducer.

Transducer. A transducer should be capable of converting the biorecognition event into a measurable signal .Typically, this is done by measuring the change that occur in the bioreceptor reaction. For example, the enzyme glucose oxidase (used as a bioreceptor in a glucose biosensor) catalyzes the following reaction:Glucose + O2 --------> Gluconic acid + H2O2

 To measure the glucose concentration, 3 different transducers can be used: An oxygen sensor that measures oxygen concentration A pH sensor that measures the acid (gluconic acid) production A peroxide sensor that measures H2O2 concentration. Note: An oxygen sensor is a transducer that converts oxygen concentration into electrical current. A pH sensor is a transducer that converts pH change into voltage change. A peroxidase sensor is a transducer that converts peroxidase concentration into an electrical current.

The choice of the biological recognition element is the crucial decision that is taken when developing a novel biosensor design.

It is important to define criteria for, for example, a suitable redox enzyme for a specific biosensor.

Most importantly, the enzyme needs to selectively react with the analyte of interest.

The redox potential of the primary redox center needs to be within a suitable potential window (usually between − 0.6 and 0.9 V vs. Ag/AgCl).

The enzyme needs to be stable under the operation and storage conditions of the biosensor and should provide a reasonable long - term stability.

Advantages and Disadvantages of using Enzymes in sensors

ADVANTAGES DISADVANTAGES

• Highly selective

• Catalytically active

improve sensitivity

• Fairly fast-acting

• One of the most known

biological components

• Expensive

• A loss activity when

immobilized on a

transducer

• Tending to lose activity

after a relatively short

period time

Considerations for biosensor development

Selection of a suitable biorecognation molecule

Selection of a suitable immobilization method

Selection of a suitable transducer

Designing of biosensor considering measurement range, linearity, and minimization of interference

Packaging of biosensor

Biosensors are classified according to the parameter thatis measured by the physicochemical transducer of the biological event as: Optical, Electrochemical ---- > Amperometric Acoustic, Thermal.

Amperometric Biosensors: the oldest ones, which have led to the higher number of ready to- use devices, are based on the monitoring of electron-transfer processes

Redox (reduction-oxidation) reactions include all chemical reactions in which atoms have their oxidation state changed. 

Amperometric Biosensors produce a current proportional to the concentration of the substance to be detected.

The most common amperometric biosensors use the Clark Oxygen electrode.

In the glucose Amperometric Biosensor, the Clark Oxygen electrode is separated from glucose by a membrane, that is permeable to oxygen. A biocatalyst Glucose Oxidase(GOD) is housed between this membrane and another membrane that separates it from the glucose. This membrane that separates GOD and glucose is permeable to both Oxygen and Glucose.

In effect, the enzyme GOD is immobilized between two membranes, the top being permeable only to oxygen and the bottom to both Oxygen and Glucose.

The Glucose that enters the membrane is Oxidised in presence of the enzyme GOD, to produce Glucuronic acid and Hydrogen Peroxide.

glucose + O2 – >  glucuronic acid + H2O2

Hence the concentration of oxygen decreases as it moves up through the membranes to reach the cathode. This decrease in Oxygen concentration is reflected as a decrease in current between the electrodes.

Alternatively, the decrease in the concentration of Hydrogen Peroxide can also be used to find the concentration of glucose, by changing the voltage applied between the electrodes to +0.68 V relative to the Ag/AgCl electrode causing the reactions:

Pt anode     H2O2 -- > O2 + 2H+ + 2e-

Ag cathode     2AgCl + 2e- -- > 2Ag0 + 2Cl-

1st generation: the normal product of the reaction diffuses to the transducer and causes electrical response

1st generation: the normal product of the reaction diffuses to the transducer and causes electrical response

1st generation: the normal product of the reaction diffuses to the transducer and causes electrical response

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

2nd generation: involves specific mediators between reaction and transducer to generate improved response

2nd generation: involves specific mediators between reaction and transducer to generate improved response

2nd generation: involves specific mediators between reaction and transducer to generate improved response

2nd generation: involves specific mediators between reaction and transducer to generate improved response

2nd generation: involves specific mediators between reaction and transducer to generate improved response

3rd generation: reaction itself causes the response

3rd generation: reaction itself causes the response

AMPEROMETRIC BIOSENSORS

Substrate Bioreceptor Product detected

Range, mM

choline choline oxidase H2O2 500ethanol alcohol oxidase H2O2 0 - 10formaldehyde f. dehydrogenase NADH 10-3glucose glucose oxidase H2O2, O2 0-7 g/Lglutamine glutamine oxidase H2O2 0-25glycerol g. dehydrogenase NADH, O2

hypoxanthine x. oxidase H2O2 4-180lactate lactate oxidase H2O2 1-40

oligosaccharidesglucoamylase, glucose oxidase H2O2 0.1-2.5

phenol polyphenol oxidasequinone

PERFORMANCE FACTORS

Selectivity Sensitivity Accuracy Response time Recovery time Lifetime

PERFORMANCE FACTORS

References A.G. Elie,Principles of Potentiometric and Amperometric

Biosensors,University of Virginia,(2002) P.V. Climent, M.L.M.Serralheiro, and M.J.F. Rebelo, Pure and Applied

Chemistry 73, pp.1993-1999, 2001 http://www.lsbu.ac.uk.html http://www.gatewaycoalition.org/files/Hidden/sensr/tocsenf.htm http://techramble.wordpress.com/2009/08/03/amperometric-biosensors/ http://www.lbb.ethz.ch/Education/Biosensors/2011FSlesson4TZ

Thank You