Clinical Biosensors: An interfacing Challenge and a Materials Question

Pankaj VadgamaInterdisciplinary Research Centre (IRC) in Biomedical Materials

Queen Mary, University of London

So how much more chemistry do we really need?

IRC in Biomedical Materials

Founded 1992EPSRC Core Grant Focus on BiomaterialsCross-disciplinary cultureMaterials, Engineering, Dentistry17 senior staff

Why biosensors?

Direct transduction(Bio)selectivitySimple, monolithic structuresMiniaturisedElectrical/optoelectronic readoutContinuous monitoringDeskilled usein vivo / ex vivo / in vitroPOCTTissue + blood monitoring

User advantages

Consumer commodityMedical ‘bypass’CheapReliableNo sample preparationDisposable Clean technology

The right chemistry ?

Ionic Substitution

Si-HA

Bone ingrowth (3 week timepoint)

HA

1 mm 1 mm

HA Purity

HA + CaO50 µm 50 µm

Pure HA

Surface modification of biosensors

Modification of materials interfacial properties in contact with biofluids in order to:

• Create a selective barrier• Allow transport of targeted analyte• Reduce fouling and maintain performance• Improve long term biocompatibility

(inflammation/coagulation)

Organic polymer membranes

• Cellulose acetate

• Poly(Vinyl Chloride)

• Nafion®

Self-assembled monolayers

• Alkyl thiols

• Organosilanes

Materials used

Polymer membranes for biosensor interfacing

Classification of polymer membranes:

• Polymeric constituents

• Structural anisotropy

• Pore size(Provides aperture control on biosensors)

Membrane technology

Particles, cells (≥ 50 nm)

MICROFILTRATION

Macromolecules, colloids (≥ 5 nm)

ULTRAFILTRATION

Organics of high M.W. (≥ 0.5 nm)

REVERSE OSMOSIS

Calibration of lactate enzyme electrode with outer, cast PVC membranes incorporating different

amounts of Triton X-100

90% 45%

27%

18%

Formation of poly (phenol) on Pt electrode(5mM phenol, pH7.4, 50mV/s)

Effect of whole blood on (a) bare (b) poly (phenol)

(a)

(b)

Needle electrode

Open microflow

0 80 120 160 200 280 320 360 400

Glucose (mM)

Glucose (mM)

Implantation Time (m)Implantation Time (m)

14

12

10

8

4

2

0

6

24040

G G I G

Continuous in-vivo glucose monitoring using isotonic phosphate buffer (pH 7.4) microflowContinuous in-vivo glucose monitoring using isotonic phosphate buffer (pH 7.4) microflow

Tissue glucoseBlood glucose

G = Glucose infusion I = Insulin infusion

G = Glucose infusion I = Insulin infusion

y = 0.75x + 0.33r2 = 0.23 n=15

0

2

4

6

8

0 2 4 6

blood lactate (mM)

estim

ated

tiss

ue la

ctat

e (m

M)

y = 0.79x + 0.59r2 = 0.31 n=33

0

1

2

3

4

5

6

0 2 4 6

blood lactate (mM)es

timat

ed ti

ssue

lact

ate

(mM

)

without microflow11 electrodes

with microflow5 electrodes

Blood-tissue lactate correlation

0

20

40

60

80

100

120

0 0.5 1 1.5 2 2.5 3

1.3mm6.5mm11mm17mm22mm

0

20

40

60

80

100

120

0 0.5 1 1.5 2 2.5 3

1.3mm6.5mm11mm17mm22mm

Flow rate = 300 µl/min

Flow rate = 500 µl/min

Detection of various glucose concentrations in the indirect stream with 1mg/ml GOx in the direct stream

glucose

Working electrode

GOx

0

0.01

0.02

0.03

0.04

0.05

0.06

0.07

buffer control 0.5 1 5 10 20

Glucose concentration (mM)

Elec

troc

hem

ical

cur

rent

(µA

)

Organic – aqueous interface for polymer formation

Smoother entry angle

Absence of over-and backpressure

Thin, continuous membranes

Entry anglefavours attachment

Single Y channel

Interfacial protein crosslinking20% (w/v) BSA (in buffer solution)

4% (w/v) Terephthaloyl chloride (in xylene)

Flow rates:

1000 µl/min (xylene phase)

300 µl/min (aqueous phase)

Channelwalls

Crosslinkedalbumin

membrane

Poly(pyrrole)

N

N

H

N

H

N

H H

n

Two electrode impedance

• TWO ELECTRODE; Symmetrical, Accurate Instrumentation.

• Interdigitated Electrodes (IDE)

• Conducting Polymer as Reference- 20mV AC potential

15 microns

15 microns

Thin Films Left and Thick Films (Right)

PPy/Cl

PPy/PVS

PPy/X

PPy/Col

Cell growth on PPy films

Examples of stained SVK14 keratinocytes on various substrates after 5 Days in culture (× 600)

From Growth Assays (AlamarBlue™ confirmed with Total Protein and ATP Quantitation) as well as Staining for Proliferation (PCNA), differentiation (K10) and Hyperproliferation (K16) Markers, Keratinocytes Growth was preferential on PPy substrates (PPy-Dermatan in particular) compared to bare gold

Sensing cells on gold

0

2500

5000

0 2500 5000

0

500

1000

0 500 1000

A

B

Representative Bode Plots (Left) and Complex Plane Plots (Right)of SVK14 Keratinocytes on Gold digit-Coated PC Coverslips

(A) Controls and (B) 4 × 105 Cells Seeded

Day 0 Day 7Day 1 Day 2 Day 3 Day 4

Equivalent circuit analysis

S

R

Applied Current I

Intracellular Fluid

Cellular Membrane, CIntercellular Fluid

A

RS

C

R∞ = RS / (R+S)

Rdif = R – R∞Cnew = C × KK = (R+S)2 / R2

B

(A) Schematic of Cells in Tissue and Equivalent Electrical Components, (B) Equivalent Circuit for Tissue Model and Circuit of Equal Frequency Response Showing

Relationship to (C) Cole Equation Parameters on Complex Plane RepresentationAdapted from Waterworth (2000), PhD Thesis, University of Sheffield

0

0.2

0.4

0 0.2 0.4 0.6 0.8 1.0

f = fc

f = DCf = ∞

R∞

C

Glaucoma Sensor

Retina Implant

Cochlear Implant

Sphincter sensor

Intracranial Pressure Sensor

IMU for Human Body Motion

Functional Electrical Stimulation

Gyro

Accelerometer

What next?

Multilayer membrane constructsReactive surfacesCharge / functional group controlMembrane miniaturisation for MEMSBiomimetic surfaces

Biomaterials

Interrogating signalOptical/acoustic/ultrasound/RF/microwave/inductive

Return signal

Sensor/implant combination or Implanted sensor

Signal detection and processing

Signal transmitter

BODY

Accelerating FactorsImproved technology diffusion

Reset national prioritiesBasic science advances

Economy of scale up

Regulatory BarriersSafety thresholds lower

Multinational bodiesSensational reportEthical changes

Competing technologiesNew therapeutics

Existing biomaterialsMEMS devices

Microfabrication advances

Retarding FactorsMedical conservation

Insufficient cost-benefitSocietal resistance to

technologyApplication complexity

Healthcare needsHigher patient expectationsNew surgical techniques Reduced bed occupancy

Improved diagnosticsOver the counter diagnosticsExploitation of New Biology

CostsDisproportionate increase.

Large work force requirementExtended development time

Expensive QA

New materials innovations

Obligatory

Comprehensive change inpractice

Genomic smart card

Smart Diagnostics

Chemical & Structural Biomimicry

Targeted drug therapy

Microbial resistance

Neurosensoryprostheses

Adjustable stents and catheters

Self diagnostic materials

Smart surgical tools

Sector Impact

Single specialty

No requirement Preferable

Multiple specialty

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