Biosensors – Part two
“Second part” : Two successful technologies
• Electrochemically based glucose monitors
• Biospecific interaction analysis with SPR
“Forgotten items”
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•
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Microcantilevers
Molecular imprints
Nanoparticles (Au, silica,..)
Nanotubes
Graphene
Ion sensitive field effect devices
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Glucose Sensors for Diabetes
Material provided by prof. Anthony Turner
Biosensors and Bioelectronics Centre
Department of Physics, Chemistry and Biology
Linköping University
The Diabetes Health Care Space
$0.35B
$5.6B
$5.2B
$12.1B
IMS and EAC data - 2004
Insulin
Oral Anti-Diabetics
Blood Glucose Monitoring
Lancets and other paraphernalia
Source: Frost & Sullivan
A brief chronology of home testing for glucose
Urine testing using, for
example, Clinitest Reagent
Tablets (1941)
followed by visually read paper
test strips for urine (1956)
Visually read paper strip for
blood glucose (1964)
Instrument to measure paper
strip by reflectance of light
(1969)
First electrochemical home
blood glucose monitor (1987)
First devices to fill by capillary
action: Medisense QID (1995);
Bayer /
KDK / Menarini (1996)
Yellow Springs Instrument Company Inc (YSI)
The original YSI serum-glucose
biosensor for diabetes clinics
1975
Glucose Biosensor 1975
Clark, LC & Lyons, C (1962). Annals New York
Ohio
1987
Academy ofYSI,
Sciences
102, 29.
1987
The YSI Analyser
1500 Sport Analysers
7
A conventional enzyme electrode
Enzyme Electrode Reactions
9
Mediated Enzyme Electrode
Glucose oxidase or PQQ Glucose Dehydrogenase
Cass, A.E.G., Davis, G., Francis, G.D., Hill, H.A.O., Aston, W.J., Higgins, I.J., Plotkin, E.V., Scott, L.D.L. and
Turner, A.P.F. (1984) Ferrocene-mediated enzyme electrode for amperometric determination of glucose.
Analytical Chemistry 56, 667-671.
10
Key bioelectrochemical reactions
GOx
D-glucose + H2O + O2
H2O2
Anode
D-glucose + 2 Medox+
(NH2)2CO + 2H2O +
2H+ + O2 + 2e-
GOx
Urease
+
H
C2H5OH + NAD+
NADH
Anode
gluconic acid + H2O2
gluconic acid + 2 Medred
HCO3- + 2NH4+
ADH
2NH3 + 2H+
C2H5O + NADH
NAD+ + 2e- + H+
Glucose response (mediated amperometric
electrodes)
“Too little enzyme”or
“Mediators”
Clinically Important Enzyme Electrodes
Electrode
Enzymes
Amperometric
Oxygen electrode, hydrogen peroxide detection at
platinum or carbon electrodes or mediated
amperometry
Oxidases e.g. Glucose oxidase (GOx), Lactate
oxidase, Galactose oxidase, Pyruvate oxidase, LAmino Acid oxidase, Alcohol oxidase. Oxalate
oxidase, Cholesterol oxidase, Xanthine oxidase,
Uricase.
Platinum, carbon, chemically-modified, mediated
amperometric electrodes
Dehydrogenases e.g. Alcohol dehydrogenase,
Glucose dehydrogenase (NAD and PQQ), Lactate
dehydrogenase
Potentiometric
Ammonia Gas-Sensing Potentiometric Electrode,
Iridium Metal Oxide semiconductor probe
Creatinase, Adenosine deaminase
pH Electrode, Filed-effect Transistor (FET)
Penicillinase, Urease, Acetylcholinesterase, GOx
Carbon Dioxide Gas Sensor
Uricase, inhibition of dihydrofolate reductase,
salicylate hydroxylase
Mediated Amperometric Glucose Sensors
MediSense ExacTech™ 1987:
Ferocene
Johnson & Johnson Lifescan
FastTake™ 1998:
Hexacyanoferrate
14
Mass Production: Screen Printing
Current Paradigm of Blood
Glucose Monitoring
Dispose of materials
Read test strip
Deposit blood drop
on to test strip &
insert strip
Prick finger or
arm
Load lancet into
launcher and
reassemble launcher
1-2 Minutes
Burdensome and lengthy
process is impediment to
high compliance
Integ LifeGuide – Minimally Invasive
•
Acquired by Inverness Medical Oct
2000
• Draws a tiny sample of interstial fluid
(about 1 μl) from the outermost layers
of the skin on the forearm in 8-10 secs
• The unit then analyzes it for glucose
in 30 secs
• “Key” (white section) is disposable
• Process avoids capillaries and nerve
endings therefore is bloodless and
eliminates pain associated with lancing
a finger
17
The Future for Integration
Metrika HbA1c:
Microelectronics, optics & dry
reagent chemistry inside a
self-contained, integrated,
Wen Qiao (University of
single-use device.
California) microfluidic chip
powered with a
commercially available radio
frequency transmitter for
electrophoresis
Printed RFID
trechnology
Skin RFID tattoo
The Arrival of Continuous Glucose Monitoring
(CGM)
Medtronic
Guardian
Dexcom
STS
Abbott Freestyle
Navigator
Meter Kit
Sensors/m
FDA
approval
$1,339
$350 (10x3day)
Aug 2005
$800
$240 (4x7day)
March 2006
$960-1,040
$360-390 (6x5 day)
March 2008 (CE June 07)
Reading
Frequency
1 per 5min (2h run in)
1 per 5min (2h)
1 per min (10h run in)
Reading must be checked by finger-stick method before adjusting insulin
Minimally-Invasive and Non-Invasive Systems
Pendragon Pendra
Pendra was CE approved in May 2003 and was available on the Dutch
direct-to-consumer market. A post-marketing reliability study was performed
in six type 1 diabetes patients. Mean absolute difference between Pendra
glucose values and values obtained through self-monitoring of blood
glucose was 52% and a Clarke error grid showed 4.3% of the Pendra
readings in the potentially dangerous zone E. Pendragon now bankrupt.
Cygnus Glucowatch Biographer
Cygnus Inc. in Redwood City, California, has gone
out of business and has stopped manufacturing its
meters. It sold essentially all of its assets to Animas
Corp. (which makes insulin pumps) for $10 million.
Futrex Inc 1992: Non-invasive glucose monitoring
using NIR
The U.S. Securities and Exchange Commission
charged Futrex with fraud, claiming that the Dream
Beam never worked.
21
Non-invasive Monitoring
A selection of the apparently most active from >95 companies identified. Bold = in clinical trials
Glucotrack: ultrasound +
thermal and electromagnetic
conductivity
GlucoLight
HypoMon:
4 electrodes;
electrophys
changes
Cnoga
“The science
fiction you
were speaking
about is reality
“
COMPANY
TECHNOLOGY
SITE
BioTex Inc, TX, USA
Near-infrared
Skin
Sensys Medical (Sensys GTS), AZ, USA
Near-infrared
Skin
Cascade Metrix Inc, IN, USA
Mid-infrared/microfluid
Skin
Light Touch Medical Inc, PA, USA
Raman spectroscopy
Finger
Integrity applications (GlucoTrack),
Israel
Photoacoustic spectroscopy
Ear lobe
VeraLight Inc (Scout DS), NM, USA
Fluorescence spectroscopy
Skin
Lein applied diagnostics, UK
Optical
Eye
Glucolight Corp (Sentris -100), PA, USA
Optical coherence tomography
Skin
Echo Therapeutics (Symphony tCGS, MA,
USA
Sonophoresis
Skin
Calisto Medical (Glucoband), TX, USA
Bio-Electromagnetic Resonance
Wrist
AiMedics (HypoMon), Australia
Electro-physiological
Chest skin
Biosign technologies (UFIT TEN-20),
Canada
Electro-physiological
Wrist
Cnoga Inc. (SoftTouch), Israel
Optical (cell colour distribution)
Skin
EyeSense, Germany
Bio-chemical/fluorescence
Eye (ISF)
VivoMedical, CA, USA
Sweat analysis
Skin
The Evolution of Home Blood Glucose Monitoring
Past
The original
Miles
Glucometer
Evolution of Blood Glucose Monitor products
Present
MediSense
Mediated
sensor
CGM
Medtronic
Guardian
Non-invasive
monitoring
(research)
Future
Supramolecular
Devices?
Newman, J.D. and Turner, A.P.F. (2008) Historical perspective of biosensor and biochip development. In: Handbook of Biosensors and Biochips (Eds R. Marks, D.
Cullen, I. Karube, C. Lowe and H. Weetall) John Wiley & Sons. ISBN 978-0-470-01905-4
Conclusions
Conclusions
• Mediated amperometric glucose biosensors continue to dominate the home
diabetes diagnostics market
• Peroxide based amperometric glucose biosensors predominate in the
decentralised and in vivo markets
• The sector is typified by companies seeking to acquire a full set of technologies
and pursuing high levels of integration (multi-sensors + multi-lancing &/or insulin
injection) and sophisticated data treatment, displays and transmission
• Implantable sensors are in the market and home-use automated systems coupled
to insulin infusion have been announced
• Non-invasive techniques have obvious attractions, but are meeting serious
(insurmountable?) technical hurdles
BIOSENSING WITH SURFACE PLASMON RESONANCE
”Real – time biospecific interaction analysis without labels”
Based partly on material from Dr. Stefan Löfås,
GE Healthcare, Uppsala
Biosensors and Bioelectronics Centre
Surface Plasmon Resonance (SPR) principle
Increasing n
Metal: Au ~50 nm
Wavelength λ ~700 nm
Light intensity decay length ~100 nm
k = 2π/λ
The Prelude: 25 years of SPR
Angular scanning
IgG
a-IgG
Ag
Dsp
Liedberg, Nylander and Lundström.
Sensors & Actuators,
1983, 4, 299-304,
Liedberg, B., Nylander C. & Lundstr6m, I. (1983).
Surface plasmon resonance for gas detection and biosensing. Sensors
and Actuators 4, 299-304
CM-Dextran Modified Surface,
e.g. Sensor Chip CM5
Carboxylated
Dextran
Gold film
Glass substrate
50 nm
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Hydrophilic
Non-crosslinked
Flexible
Extends from the surface
Low non-specific binding
High binding capacity
Easy to activate and use
for covalent coupling
Covalent coupling chemistries
O
Ligand
O
PDEA
COOH
EDC/
NHS
NH2
Amine
SS
Ligand
Ligand
thiol
SS
Ligand
Surface
thiol
O
C N
H
O
C NH Ligand
S
Ligand
S
SH
N
C N
H
C O N
1) Cystamine
2) DTE (DTT)
Hydrazine
O
C N
H
Ligand SS
N
O
C N
H
1) Ligand C
O
C
SH
O
H
NHNH2
2) NaBH3CN
O
C NHNHCH2 Ligand Aldehyde
Carboxyl group
for covalent
attachment of
molecules to the
sensor surface
Dextran matrix
Gold
Glass support
Pneumatic valve
Determination of
the position of
the reflectance
minimum
replaced by the
movement of a
dark band on a
diode array or
(CCD) camera
Biomolecular interactions
DNA
hybridization
Early example of BIA
The Biology Project
The University of Arizona
Identification of the subclass of an unknown monoclonal ab
Principles of biosensing with an extended coupling matrix and surface plasmon
resonance, Sensors and Actuators 8, 11 (1993) 63- 72
B. Liedberg and I. Lundström Laboratory of Applied Physics, Linköping University, S-581 83 Linköping
(Sweden), E. Stenberg, Pharmacia Biosensor AB, S-751 82 Uppsala (Sweden)
Some numbers
• SAM layer ~ 2 ng/mm2
• Dextran layer ~2-3 ng/mm2
• Surface chemistry modification and immobilization
reproducibility: typically CV< 5 %
• 1000 RU corresponds to ∆Θsp ≈ 0.1₀
• Sensitivity limits (Biacore A100)
• 1 RU ~1 pg/mm2
• Detection spot in Biacore A100 = 0.01 mm2 => 10 fg/spot
• Protein (100 kDa) binding => 0.01 attomole/spot
• Drug molecule (500 Da) binding => 2 attomole/spot
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Antigen – antibody
DNA hybridization
Drug – receptor
Pollutant – ligand
(protein, aptamer, DNA
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Antibody selection
Drug discovery; drug testing
Environmental monitoring
Food quality
Cornerstones of Biacore™ systems
Surface
Plasmon
Resonance
(SPR) detection
system
User-friendly software
Sensor chip
Microfluidic system
The Early Story of SPR for Biosensing
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1983 Liedberg, Nylander, Lundström, Surface Plasmon Resonance for gas
detection and biosensing, Sensors and Actuators 4(1983) 299-304
1984 Research program initiated within Pharmacia AB and Pharmacia
Biosensor established
1986 Fundamental principles verified in prototype
1988 Research phase concluded, product development phase started
1989 Marketing organization established
1990 Launch in October and first sale of instrument
Business aspects of the Biacore development
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1990-1993 Market introduction; survival or closed shop?
1994-1997 Market acceptance; company stabilization
1996 Pharmacia Biosensor becomes Biacore AB
1998-2002 Rapid organic expansion, brand established
2003-2004 Regression …
2005
Turn around and new growth strategy, revenues back on track
(~ 85 million USD)
2006
Biacore acquired by GE Healthcare for ~450 million USD
Unique Biological Information
Specificity
Kinetics & Affinity
Concentration
Thermodynamics
How specific is the
interaction?
How fast and how strong is
the interaction?
How much
biologically active
protein is present?
What drives the
interaction?
Data generated gives critical insights into protein
functionality, the role of proteins in normal and diseased
states, and the influence of potential drug candidates
Kinetic properties have therapeutic
consequences
– Kinetic properties affect multiple aspects of drug
function (e.g. pharmacokinetics, dosing)
Rapid kinetics: frequent administration of low dose required
to occupy target
100 % blocked target
Slower kinetics: administration of high dose
occupies target for long time
0
0
2
4
6
8
hours
This information is critical to therapeutic
performance of drugs since duration of efficacy depends on
the receptor-drug complex association and dissociation rate
constants
Use of label free kinetics for antibody analysis
Life science
research
Antibody
development
RU
14
Step 1
12
10
8
Response
13°C
6
4
2
0
wt
mutant
45°C
-2
-100
0
100
200
300
400
500
600
700
800
900
s
Time
RU
Step 2
16
11
Response
0
6
• improved affinities
1
-4
-100
100
200
300
400
500
600
700
800
900
s
Time
20
15
Step 3
Response
Biological relevance
Structure –function
Interactomics
0
RU
10
5
0
-5
-100
0
100
200
300
400
Time
500
600
700
800
900
s
• similar affinities
• slower off rates
Screen Select Characterize Improve
Industrial process
Antigen and Fc Receptor binding
Possibilities
• Imaging (S)PR (multispot arrays)
• Plasmonics in nanostructures/-particles
• Combination of the two
• SPR and mass spectrometry
• SPR and QCM - D
• SPR and microcalorimetry
An Optofluidic Nanoplasmonic Biosensor for Direct
Detection of Live Viruses from Biological Media
Yanik et. al., Nano Lett. 10 (2010) 4962 – 4969
Nanostructured digital microfluidics for enhanced surface
plasmon resonance imaging,
Malic, Veres, Tabrizian, Biosensors & Bioelectronics 26 (2011) 2053 - 2059
Conclusions
• Biosensors have achieved considerable success in both the commercial and academic
arenas and the need for new, easy-to-use, home and decentralised diagnostics is greater
than ever
• Amperometric biosensors continue to dominate the market
• Electrochemical biosensors are available for a range of decentralised analyses,
including medical, food and environmental applications
• Array-based fluorescence sensors are now well established for genomic and proteomic
assays
• Label-free assays based on SPR dominate the laboratory affinity sensor market
especially in drug development
• The search continues for a new disruptive technology!
Electrochemistry in Diabetes Management
Adam Heller
ACCOUNTS OF CHEMICAL RESEARCH, Vol. 43,
No. 7 July 2010 963-973
Direct optical detection in bioanalysis: an
update
Günter Gauglitz
Anal Bioanal Chem (2010) 398:2363–2372
A survey of the 2006–2009 quartz crystal
microbalance biosensor literature
Bernd Becker and Matthew A. Cooper
J. Mol. Recognit. 2011; 24: 754–787
Lab-on-a-chip based immunosensor principles
and technologies for the
detection of cardiac biomarkers: a review
Mazher-Iqbal Mohammed* and Marc P. Y.
Desmulliez
Lab Chip, 2011, 11, 569–595