MEMS based sensors for cellular studies

Taher SaifMechanical Science and Engineering

University of Illinois at Urbana-Champaign

Part of GEM4 Summer School lectures on instruments for cell mechanics studies (Aug 10, 2006, MIT)

Anchoragesite

Actin filaments

Develop portable micro sensors to study:

• Cell mechanical response

• Cell adhesion

in different biochemical environmentsto explore mechanotransduction and disease detection

U. of Illinois at Urbana-Champaign

Objective

F

FF

F

Cell membrane

Basic idea

U. of Illinois at Urbana-Champaign

Functionalized probe

A micro spring is used to measure cell force

actin

Cell membrane

U. of Illinois at Urbana-Champaign

Basic idea

Functionalized probe contacts a cell and formsadhesion site

actin

Cell membrane

U. of Illinois at Urbana-Champaign

Basic idea

Probe is moved away from the cell. The cell appliesa force on the spring. The force is measured from the spring deformation and its spring constant.

The cell may also be compressed or indented.

P dK (spring constant ) = 3EI/L3

P = Kd

Typical K ~ 10 nN/µm

Calibration: 1) Resonant frequency, geometry, elastic property2) Comparing with another spring (e.g., AFM)

Cantilever as a mechanical spring

L

I=moment of inertia = width x depth3/12

Simple implementation

U. of Illinois at Urbana-Champaign

Lδ

Κ= 3EI /L3F=Kδ

δ

Cell attachedto substrate

F = cell force = Kδ• Force sensor (such as a cantilever) is coated by fibronectin

• It is calibrated to determine spring constant, K

• The sensor tip is brought in contact with the cell - focaladhesion sites form

• It is moved away from the cell. The cell force is measured fromthe deformation δ

Advantages: - Force sensor independent ly calibrated- Force is applied at an anchorage site- In-situ observat ion

U. of Illinois at Urbana-Champaign

Experimental setup 1 µmResolut ion

20nm resolut ionpiezo actuator

xY

z

cell

MEMS cant ileverCell culturemedium SCS chip

Microscope object ive

5o

5oX-Y-Zst age

X

ZY

cell

5 deg5o

Stage motionMEMS cant ilever

U. of Illinois at Urbana-Champaign

MEMS force sensor: beams anchored at both ends

F = Kx

xSisubstrate

K = 3.4 nN/µm

Cell

Probe

Probe

Flexural springs (1µm wide)

Yang and Saif. Review of Scientific Instruments 76, 044301 (2005).

Example

2D force sensorExample

U. of Illinois at Urbana-Champaign

Endothelial cells in a culture dish with MEMS cantilever

Bovine endothelialcells on a substrate

MEMS cantilevercoated with fibronectin (K=18nN/µm)

100µm

The MEMS cantilever has formed adhesivecontact with the cell

5 deg3-5 deg

Schematic of theMEMS cantilever

Invertedmicroscope

cell

Example

U. of Illinois at Urbana-Champaign

Force response of an endothelial cell

Microtubule

C0

102030405060708090

100

0 20 40 60 80 100Distance (µm) of the cantilever end from a fixed reference

B1D1

C1

120

1

2 34

5

OCell deformation

Cell deformationbegins

A1

Necking

Motion

A1 B1Endothelialcell

C1

D

D1

Just afterfailure

Relaxedcell

g

U. of Illinois at Urbana-Champaign

Force response of an endothelial cell

Microtubule

C0

102030405060708090

100

0 20 40 60 80 100Distance (µm) of the cantilever end from a fixed reference

B1D1

C1

120

1

2 34

5

OCell deformation

Cell deformationbegins

A1

Necking

Motion

A1 B1Endothelialcell

C1

D

D1

Just afterfailure

Relaxedcell

g

U. of Illinois at Urbana-Champaign

Force response of a monkey kidney fibroblast cell

-20

0

20

40

60

80

100

120

0 10 20 30 40 50 60 70 80 90

Cell deformation (µm)

ForwardBackward

Forc

e (n

N)

Yang and Saif. Experimental Cell Research 305 (2005) 42– 50.

Cell force response under stretch is linear and reversible

-10

0

10

20

30

40

50

60

0 5 10 15 20 25 30 35 40Cell stretching (µm)

Stretching-1Un-stretching-1Stretching-2Un-stretching-2Strecthing-3Un-strecthing-3Before

CytoD

After CytoD

Monkey kidney fibroblast

U. of Illi ois at Urbana-Champaign

Cyto-D treatment disrupts force bearing capacity

Yang and Saif. Experimental Cell Research 305 (2005) 42– 50.

Cyto-D disruptsactin network

U. of Illinois at Urbana-Champaign

Force response of a monkey kidney fibroblast cell

-15

-10

-5

0

5

10

15

20

25

30

-45 -40 -35 -30 -25 -20 -15 -10 -5 0 5 10 15 20 25 30

Cell deformation (µm)

Push-1Pull-1Push-2Pull-2

(1)(2)

(3)

(4)Fo

rce

(nN

)

Tension

Indentation

Cell response under indentation is non-linear and hysteretic

-20

0

20

40

60

80

100

120

140

0 5 10 25 30 35 40

Indentation

15 20

Mechanism of non-linearity and irreversibility under indentation

GFP actin

Force measured from change of gap betweentwo markers

Sl=2.8, R ~ tα, α=2.8

α=.7

Log(

dia)

Log(time)

Actin agglomerates irreversibly under indentation

Yang and SaifActabiomaterialia 2006 (in press)

44:1848:00 71:42

Monkey kidney fibroblast subjected to mechanical indentation (injury simulation). Here actin stress fibersare highlighted by green florescent protein (GFP). In response to indentation, the cell signals localactin agglomeration at discrete locations. Such actin agglomeration is also observed in various physiologicalconditions such as during ischemic attack in kidney cells. This is the first evidence of actin agglomeration dueto mechanical stimulus (Shengyuan and Saif, Actabiomaterialia 2006, in press).

38:42

45 µm

probe

More evidence of actin agglomeration

Actin agglomeration in physiological condition: ischemic attack

Ashworth et al. Am J. Physiol Renal Physiol 284: F852, 2003.

Porcine kidney cells

Why MEMS bio sensors:

1. Force range: 1-100 nN (natural progressionfrom optical tweezer, magnetic beads, AFM)

1. Flexibility of design (cell contact region may be designed in a variety of fashions)

3. Large cell deformation range (sub µm-10s of µm)