BIOSENSORS (1) industrial microbiologyppt

Tried to explain it in a simple way

1. Block diagram of biosensor
A biosensor is an analytical device which is used to
determinethe presence and concentration of a specific
substance in a biological analyte
Biosensor
DisplayBioreceptor Transducer
Signal
Processing
Desired molecule
Introductionto Biosensors
Biosample

2. Basiccharacteristics of a biosensor
LINEARITY:Maximumlinearvalueofthesensorcalibrationcurve.
Linearityofthesensormustbehighforthedetectionof
highsubstrateconcentration.
SENSITIVITY: The value of the electrode response per substrate
concentration.
SELECTIVITY: Interference of chemicals must be minimized for obtaining
the correct result.
RESPONSE TIME: The necessary time for having 95% of the response.

3. Introduction to Biosensors
Bioreceptor Transducer
Antibody
Enzyme
Nucleic Acid (DNA)
Cell
MIP
Optical
Electrochemical
Mass based
Temperature based
potentiometric
amperometric
conductimetric
Electric&
Magnetic
Dielectric properties
Permeability properties
Voltage or Current
Fluorescence
Interference
Absorption

4. Applicationsof biosensors
Glucose monitoring in diabetes patients←historical market driver
Environmental applications e.g. the detection ofpesticidesand river water
contaminants such as heavy metal ions
.
Remote sensing of airbornebacteriae.g. in counter-bioterroristactivities
Detection of pathogens
.
Determining levels of toxic substances before and afterbioremediation
Detection and determining oforganophosphate
Routine analytical measurement offolic acid,biotin,vitamin
B12andpantothenic acidas an alternative tomicrobiological assay
Determination ofdrug residuesin food, such asantibioticsandgrowth
promoters, particularly meat and honey.
Drug discovery and evaluation of biological activity of new compounds.
Protein engineering in biosensors
Detection of toxic metabolites such asmycotoxins

5. Amperometricbiosenor
Amperometricbiosensorsareself-containedintegrateddevices
basedonthemeasurementofthecurrentresultingfromthe
oxidationorreductionofanelectroactivebiologicalelement
providingspecificquantitativeanalyticalinformation.

6. Blood Glucose Monitoring
What is it?
Blood Glucose Monitoring is a way of checking the
concentration of glucose in the blood using a glucometer.
What is the purpose?
Provides quick response to tell if the sugar is high or low
indicating a change in diet, exercise or insulin.
Over time, it reveals individual of blood glucose changes.

7. Why monitor blood glucose?
Reduces risk of developing complications with diabetes.
Allows diabetics to see if the insulin and other
medications they are taking are working.
Gives diabetics an idea as to how exercise and food
affect their blood sugar.
May prevent hypoglycemia or hyperglycemia

8. AmperometricGlucose Biosensor
Developed by Updike and Hicks
Enzyme Glucose oxidasecatalyze the oxidation of glucose by
molecular oxygen producing glucolactone and hydrogen peroxide.
In order to work as a catalyst, GOxrequires a redoxcofactor –flavin
adenine dinucleotide(FAD), works as an initial electron acceptor and
is reduced to FADH
2.
Glucose + GOx–FAD
+
Glucolactone + GOx–FADH
2

9. •The cofactor is regenerated by reacting with oxygen, leading to the
formation of hydrogen peroxide
GOx–FADH
2 + O
2 GOx–FAD + H
2O
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
2O
2 2H
+
+O
2 + 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
Goxto the electrode.

11. Types of glucose biosensors
Enzymaticglucose biosensors
•First generation glucose biosensor
•Second generation glucose biosensor
•Third generation glucose biosensor
Non-enzymaticglucose
biosensors

12. Generations
1
st
generation: the normal product of the reaction
diffuses to the transducer and causes electrical
response
2
nd
generation: involves specific mediators between
reaction and transducer to generate improved response
3
rd
generation: reaction itself causes the response

13. First generationglucose biosensors
Thefirstgenerationglucosebiosensorsestimatedglucoseconcentrationinthe
samplebasedonhydrogenperoxideproductionbyglucoseoxidaseutlizing
dissolvedoxygen.
•Basedonthistechnology,YellowspringInstrumentcompany,launchedthe
firstcommercialglucosebiosensorinmarketin1975forthedirect
measurementofglucose.
•Theusageofthemostexpensivemetalplatinumforthefabricationofthis
electroderestrictedthebiosensortoclinicallaboratoriesonly.

14. Major drawbacks of first generation glucose
biosensors
•Amperometricmeasurementofhydrogenperoxiderequireda
highoperatingpotential(0.6V)forhighselectivity.
•Restrictedsolubilityofoxygeninbiologicalfluids,which
producedfluctuationsintheoxygentension.
•Deactivationoftheenzymeduetotheproductionofhydrogen
peroxide.

15. Second generation glucose biosensor
•The second generation glucose biosensor utilized redox mediator to transfer
electrons from the enzyme to the working electrode surface.
•Avarietyofredoxmediators,suchasferrocene,ferricyanide,quinines,methylene
blueetcwereusedtoimprovesensorperformance.
•Usageofredoxmediatoreliminatedtheneedofoxygenforelectrontransferatthe
electrodesurface,thusovercomingthedrawbackoflimitedoxygenpressureobserved
inthefirstgenerationbiosensor.
•Thelowerredoxpotentialofchosenmediators(0-2V)resultsinnointerferencefrom
otherelectroactivespeciessuchasuricacid,ascorbicacid.
•Redoxmediatorenhancestheelectrontransferbetweentheredoxcenterofenzyme
andtheelectrodesurface.

16. Major drawbacks of second generation glucose
biosensors
•High competition between redox mediator and oxygen.
•Interference of other electroactivespecies 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.

17. Third generation glucose biosensors
•Thethirdgenerationglucosebiosensorsarebasedonthedirectelectron
transferbetweentheactivecenterofenzymeandtheelectrode.
•Theintrinsicbarriertoelectronflowistheglobularstructureofglucose
oxidasewiththeactivesite,containingFAD/FADH
2redoxcofactor,buried
deepinsideacavityof~13A
◦
isamajorhinderancefordirectelectron
transfer.
•Carbonnanotubesimmobilizedelectrodesurfaceprovidesuitableorientation
forenzymeimmobilizationandestablishconnectionbetweenelectrodesurface
anddeeplyburiedactivesiteofenzyme.

18. Non-enzymaticglucose 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 oxide
layer OH
adsthat mediate oxidation of the adsorbed species.

19. Glucose Biosensors Based on Carbon
Nanotube Nano electrode Ensembles
Thedevelopmentofglucosebiosensorsbasedoncarbonnanotube(CNT)
Nanoelectrodeensembles(NEEs)fortheselectivedetectionofglucose.
CNTshaveahighelectrocatalyticeffectandafastelectron-transferrate.
GlucoseoxidasewascovalentlyimmobilizedonCNTNEEsvia
carbodiimidechemistrybyformingamidelinkagesbetweentheiramine
residuesandcarboxylicacidgroupsontheCNTtips.
Thecatalyticreductionofhydrogenperoxideliberatedfromtheenzymatic
reactionofglucoseoxidaseupontheglucoseandoxygenonCNTNEEs
leadstotheselectivedetectionofglucose.
Thebiosensoreffectivelyperformsaselectiveelectrochemicalanalysisof
glucoseinthepresenceofcommoninterferents(e.g.,acetaminophen,
uricandascorbicacids),avoidingthegenerationofanoverlappingsignal
fromsuchinterferers.Suchanoperationeliminatestheneedfor
permselectivemembranebarriersorartificialelectronmediators,thus
greatlysimplifyingthesensordesignandfabrication.

20. •ThefabricationofglucosebiosensorsbasedonCNTNEEs
fortheselectiveandsensitivedetectionofglucose.CNT
NEEseliminatepotentialinterferencethroughthepreferential
detectionof hydrogenperoxide.Such developmentof
interference-freetransducersshouldsimplifythedesignand
fabricationofconventionalandminiaturizedsensingprobes.
TheglucosebiosensorbasedonanalignedCNTNEEisthus
suitableforthehighlyselectivedetectionofglucoseina
varietyofbiologicalfluids(e.g.,saliva,sweat,urine,and
serum).The biosensorfabricationtechnologydemonstrated
inthisworkisreadilyapplicabletothefabricationofother
biosensorsbasedon oxidases,suchasbiosensorsfor
cholesterol,alcohol,lactate,acetylcholine,choline,hypox-
anthine,andxanthine.

21. Glucose biosensor test strips
Meter
Read glucose
Dry coating of GO + Fc
Patient adds drop of blood,
then inserts slide into meter
I
t
Patient reads glucose level on meter
e’s
electrodes