Basic QCM-D and Q-Sense Product range

Malin Edvardsson, PhD

Product manager, Q-Sense

P A G E 1

O U T L I N E

Background

Basic QCM-D Theory

What can QCM-D Characterize

Application examples

Q-Sense Product Offering

P A G E 2

QCM AE QCM-D AE Q-Sense

1960s: QCM for monitoring of thin films in air and vacuum

1972 QCM as biosensor

1980s QCM is operated in liquid

1990s QCM is further developed into QCM-D

1995 QCM-D technology patented

1996 Q-Sense founded

1999 1st QCM-D generation launched

2005 2nd QCM-D generation launched

2010 Q-Sense => Biolin Scientific

2013 >500 instruments in >30 countries

>1400 publications

P A G E 3

M E A S U R E M E N T

P R I N C P L E

P A G E 4

What does QCM-D offer?

Follow molecular events in real-time, in liquid or in air Measure mass (ng) and thickness of molecular layers (resolution 1Å-1 µm) Analyze structural and mechanical properties of molecular layers

Water content info, swelling/contraction etc

Flexible choice of surfaces/samples

HOW?

P A G E 5

QCM-D: a sensor-based, acoustic technique

Top View Bottom View

electrodes

Quartz Crystal covered with Au layer

Sensor Diameter 14 mm

Acoustic= based on increases/decrease in oscillation frequencies (Quartz Crystal Microbalance with Dissipation monitoring)

P A G E 6

How does the sensor oscillate?

f = Change in frequency (Hz) tq= thickness of quartz

Vq= 3420 m/s

Overtones

n = 1 n = 3

If fundamental frequency 5MHz:

q qq t vnvnf 2

Resonance condition

~ tq Ʌ tq dtq

P A G E 7

Rigid film

large Ɏ ĺLow D

Soft film

small ɎĺHigh D

QCM - D

ǻf (ĺfilm mass)

ǻD (ĺ film viscoelastic properties)

Monitoring:

Rodahl, M. et al.; Review of Scientific Instruments 1995, 66, 3924-3930 Rodahl, M. and Kasemo, B. Review of Scientific Instruments 1996, 67, 3238-3241

D-D: Dissipation

P A G E 8

U Rodahl, M. et al.; Review of Scientific Instruments 1995, 66, 3924-3930 Rodahl, M. and Kasemo, B. Review of Scientific Instruments 1996, 67, 3238-3241 Find resonance ~ ms decay recording ~ ms data communication ~ ms

Ɏ Umax

Umax/e

t QCM-D f

Q-Sense sensor excited to resonance

Drive voltage is perodically swithced on and off

P A G E 9

Schematic QCM-D measurement

ȟfn1, ȟDn1

ȟfn2, ȟDn2

ǻD time

ǻf ȟf

ǻD ǻD

t U ~1V

I(t)=I0·e-t/Ɏ sin (2Ɋft-ɐ)

Mass coupled to the surface

viscoelasticity of the coupled layer

P A G E 10

Systems that QCM-D can characterize

Surface adsorption/desorption

Biomolecules (protein, vitamin, antibody, DNA, ...)

Polymers/polyelectrolytes

Cells

Surface reaction

Conformation change (protein, DNA, polymer, cells)

Crosslinking (protein, polymer, ...)

Hydration (polymer)

Bulk characterization

Viscoelastic properties of fluids (protein solutions, ...)

P A G E 11

QCM-D Application overview

P A G E 12

APPLICATION EXAMPLES

P A G E 13

Adsorption

P A G E 14

QCM-D in protein drug formulation and storage

Protein in contact with surfaces

(concentrators, filters, containers, syringes, tubing, beakers)

Risk of protein unfolding and aggregation

Surface interactions poorly understood

P A G E 15

Surface interactions of monoclonal

antibodies

Parameters

Two different antibodies

(one stable, one known to self associate)

High/low concentrations

(1 mg/ml, 50 mg/ml)

With/without surfactant

(PS-80)

Four surfaces

(Au, PS, teflon and silica)

QCM-D Outcome

Mass/thickness

Viscosity

Shear modulus

Oom et al - J. Pharm. Sc. - 2011

P A G E 16

Effect of surface and surfactant

mAb2 generally adsorbs

stronger than mAb1

Addition of PS-80 reduces

protein aggregation for both antibodies Adsorption of protein onto different surfaces from 1 mg/ml (low conc) solutions of mAb1 and mAb2

Oom et al - J. Pharm. Sc. - 2011

P A G E 17

Swelling

P A G E 18

Cell membrane water permeability

P A G E 19

Effect of composition on water permeability of

model stratum corneum lipid membranes

Myung Han Lee et al: Soft Matter, 2012, 1539-1546

Deposition and swelling of a supported

membrane under introduced humidity

The swelling kinetics correlates to

diffusion of water molecules

P A G E 20

Permeability (P), diffusivity (D) and solubility (S) of water vapour in the membrane as a function of membrane thickness, FFA chain length, saturation level, and CER structure

Myung Han Lee et al: Soft Matter, 2012, 1539-1546

P A G E 21

Crosslinking

P A G E 22

Adsorption

Protein adsorption and re-arrangment

P A G E 23

Protein adsorption and re-arrangment

MefP-1 mussle adhesive protein- elongated structure

NaI04- crosslinker- release of water

P A G E 24

Modeling results

before after x 1.04103 1.18103 kgm-3 x (QCM-D) 22.4 7.3 nm x (QCM-D) 1.8 x10-3 6 x10-3 Nsm-2 x (QCM-D) 6.6 x104 3 x105 Nm-2 NaIO4

Release of Water

Protein adsorption and re-arrangment

before after

P A G E 25

Degradation

P A G E 26

Follow break-up of films,

mass removal, swelling,

Three commercially

available kitchen spray cleaners treating a sensor coated with a model stain (triolein ~40 nm).

1: Stain put on

sensor, and detergent added.

2: Detergent

penetrates film, and film swells

3. Film stain

degradation starts.

Q-Sense 2008

Detergent effectiveness

P A G E 27

Polyelectrolytes & Nanoparticles

P A G E 28

Nanoparticles for drug delivery

Great potential for non-

invasive drug delivery (oral, nasal, pulmonary)

P A G E 29

Structural rearrangements of polymeric Insulin-loaded Nanoparticles Interacting with Surface-Supported Model Lipid

Membranes

Frost et al. J. Biomat. and Nanobiotech., 2011, 2, 181 - 193 (Anionic)

Polyelectrolyte

complexes

P A G E 30

Formation of a supported lipid bilayer

Mimic for human

cell membranes

P A G E 31

NP-HI interaction with model membranes of different charge

Positive membrane

Negative membrane

Neutral/ slightly

negative membrane

High dissipation = soft film

Spreading of overtones = soft film

Frost et al. J. Biomat. and Nanobiotech., 2011, 2, 181 - 193

1.Lipid membrane (pre-formed)

2.Injection of Nanoparticles

3.Rinsing

P A G E 32

Schematics of possible NP adsorption

Frost et al. J. Biomat. and Nanobiotech., 2011, 2, 181 - 193

P A G E 33

Follow-up study: Release of three NP/Insulin complexes with reducing agent (glutathione) Frost et al. J. Coll. Int. Sci. (2011) 362(2) 575-583

1.Pre-formed bilayer

POPC:POPS (3:1)

2.Three diffrerent NP

complexes (fused to insulin via S-S bond)

3.Reducing environment a

mimic for intra cellular conditions

P A G E 34

Mesuring cell attachement

P A G E 35

Reversible Changes in Cell Morphology due to Cytoskeletal rearrangements measured in Real-time by QCM-D

Tymchenko N., et al., Biointerphases, (2012) 7:43

P A G E 36

Tymchenko N., et al., Biointerphases, (2012) 7:43

Protocol

SiO2-surface, 37ºC, Flow 50µl/min

Coat surface with ECM protein

Adsorb Collagen I

Expose to serum containing medium

Seed cells in situ

A.Flow cells over surface

Fibroblasts (NIH3T3 or HS483.T)

B.Let attach...

C....and spread

Induce morphological changes

D.Add cytomorphic agent

Cytochalasin D

P A G E 37

MICROSCOPY

Tymchenko N., et al., Biointerphases, (2012) 7:43

Live cell images of HS 483, T on collagen and serum coated sensors

Cell spreading prior

to Cytochalasin D exposure

Cell retraction after

20 in exposure to

Cytochalasin D

Cell recovery after

Cytochalasin D removal

P A G E 38

Tymchenko N., et al., Biointerphases, (2012) 7:43

Different phases clearly

distinguishable via Df-plot:

A.Seeding

B.Attachment

C.Spreading

P A G E 39

Q SENSE PRODUCT OFFERING

P A G E 40

Q-SENSE PRODUCT FAMILY

Q-Sense E4 Q-Sense E1 Q-Sense Omega Auto

P A G E 41

Q-Sense E4

4 sensors

Controlled one-direction flow

High chemical compatibility

HPLC fittings

Complete software package

Sample volume ~200µl

Temp range 15-65° C

Temp stability +/- 0.02°C

Sensitivity 0.5 ng/cm2

Stability

P A G E 42

The E4 Measurement Chamber

parallel 2 by 2 parallel serial 2 by 2 serial

Examples of

Experimental Design

P A G E 43

Q-Sense E-series Modules

High

Temperature

Chamber

Humidity

Electrochemistry Window Ellipsometry

Open PTFE

ALD Holder

Standard

Flow

P A G E 44

Q-Sense E1

Same characteristics

as E4, but one module

Especially suitable for

combinations with ellipsometry and microscopy

P A G E 45

Launched 2012

Key words

Ease of Use

Integration

Automation

Save time

Reproducibility

147 kEuro

The next Generation Q-Sense Instrument

Q-Sense Omega Auto

P A G E 46

Features at a Glance

Integrated turn-key system

Automated sample handling

FlexiFlow

2 x 4 sensor module

Versatile sample tray

Waste and rinse station

4 - 70°C

30 µl/sensor

Same sensors

E1 & QHTC

P A G E 47

Q-Sense Sensors

Costum made:

Any material that can be applied

as a thin homogeneous film

Wide Array of Sensors (>30 available)

Glass, ceramics

Plastics (PP, PS)

Stainless Steel

Basic elements (e.g. Ti, Cu, Au)

Oxides (e.g. Silicon Dioxide)

Nitrides, Sulfides and Carbides (e.g. TaN, ZnS)

Hydroxy Apatite

Cellulose

Biotin functionalized

HisTag

14 mm diam.

5 MHz

P A G E 48

Summary the QCMD technique

Probes Mass and Structure/Mechanical properties

Real time

Label free/non-invasive

High resolution (~ 200 datapoints/s )

Flexibility in measurement conditions (flow, temp, etc)

Flexible choice of surface

Combine with other techniques e.g.

microscopy, electrochemistry/impedance

THANKS!

Malin Edvardsson

malin.edvardsson@biolinscientific.com +46 708 738269
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