Date post: | 14-Jul-2015 |
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Technology |
Upload: | dr-pranav-rathi |
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Susan Atlas—Lead of the DTRA projectUNM Physics / Cancer Center / Director of CARC
Haiqing Liu (G. Mantano lab)—Microdevice applications of kinesinLANL & Center for Integrated Nanotechnology (CINT)
Collaborations
Funding DTRA—DTRA CB Basic Research Program under Grant No. HDTRA1-09-1-008
Dr. Larry Herskowitz
Dr. Anthony Salvagno
Dr. Andy Maloney
Dr. Steven Koch
Acknowledgments
Outline
• Introduction to optical tweezers• Design and construction• Automation and control• Optical tweezers calibration• DNA sample preparation• Results
c
PQnF =
tpF ∆
∆=
22gs FFF +=
Optical Tweezers are used to apply forces over nanometer scale on the order of piconewtons
Design considerations
• Force ~ 65pN with .530nm (diameter) polystyrene beads
• Stability and precision
• Fast, user-friendly and automated
• Safety
Some problems with the design!
• Accessibility to optomechanical controls of Z lens, QPD and microscope
• Temperature hike inside enclosure
• Mechanical vibration nose
Accessibility problem was solved by extending optomechanical controls
Z-lens controls QPD controls
Microscope focus control
Fiber plugin adapterMicroscope inlet
Fiber feeder
Temperature hike problem was solved bydeveloping Fiber light
The parameters we calibrate!
XKF x−=
Kx is the stiffness in x direction
X is displacement of bead center from the trap center
Zb is the distance between beamwaist and the trap center.
X
Z
Trap center
Beam waist
Surface
Zb
Calibration of stiffness Kx We use Brownian noise to map the stiffness
Tkfrtxm
tftxKtxtxm
B
x
βωπηβ
β
4)(;6;0)(
)()()()(2 ===
+−−=
After Fourier transformation
πβ
ωπβ
βπω
ββωπω
2
)2
(
)(~
4)4()(~
22
2
22222
xc
x
B
Bx
Kf
K
TKx
TKKx
=
+
=
=+
Power spectrumEquation of motion for trapped bead
Cutoff frequency fc
543
161
25645
81
169
1
6
−
−
+
−
=
hr
hr
hr
hr
rπηβ
Trap center determination
At 1.2r (bead radius) from surface fc≈1/2 of bulk
• Trap center offset for big beads is 186 and small bead is 367 nm• Big bead is 1.96 times the small bead and small bead is 1.97 times farther then big bead
Corner frequency vs bead center height from surface (H2O)C
orne
r fr
eque
ncy
(Hz)
Cor
ner
freq
uenc
y (H
z)
Bead center height (multiples of r=520 nm from surface)Bead center height (multiples of r=520 nm from surface)
Corner frequency vs bead center height from surfaceH2O vs D2O
Cor
ner
freq
uenc
y (H
z)C
orne
r fr
eque
ncy
(Hz)
Bead center height (multiples of r=265 nm from surface)Bead center height (multiples of r=265 nm from surface)
Stiffness vs bead center height from surfaceH2O vs D2O
Stiffness does not depend on height but corner frequency does
Perc
eive
d st
iffn
ess
(pN
/nm
/W)
Perc
eive
d st
iffn
ess
(pN
/nm
/W)
Bead center height (multiples of r=520 nm from surface)Bead center height (multiples of r=520 nm from surface)
Estimated stiffness (H2O) .038(7) pN/nm
Average variance (H2O) 12300+/-800 mV2
Stiffness calibration results
Big beads (1.04µm; diameter)
Estimated stiffness (D2O) .04(2) pN/nm
Average variance (D2O) 12500+/800 mV2
Small beads (.530µm; diameter)
Estimated stiffness (H2O) .011(5) pN/nm
Average variance (H2O) 2100+/-200 mV2
Estimated stiffness (D2O) .012(5) pN/nm
Average variance (D2O) 2000+/-300 mV2
Calibration of detector sensitivity (DOG)
DOG scan and linear-fit on left and sensitivity extracted from slope of linear-fit vs bead position relative to beam waist on right from
DOG scans from one edge to another of bead
X sensitivity vs bead position relative to beam waist
Big beads
X S
ensi
tivi
ty (
mV
/nm
)X
Sen
siti
vity
(m
V/n
m)
Bead position relative to beam waist (nm)Bead position relative to beam waist (nm)
Sensitivity calibration results
Sensitivity of small bead is 4.5 times the big bead and the stiffness of big bead is 4.3 times the small bead
Comparison
Sensitivity for big beads at trap center 10.8+/-.5 mV/nm
Sensitivity for small beads at trap center 2.4+/-.2mV/nm
Future work
• Automate Z piezo using camera to find the surface
• Automate Z lens and QPD controls
• DNA unzipping in D2O
• Investigate DNA protein interactions in H2O and D2O
• Develop touch screen controlled automation for optical tweezers
Holder stageL4
Mirror holderLens tube holder
Dichroic holder stage platform
Top view
Clip
X
Z
YAfter reflection
X Z
YBefore reflection
Dichroic holder stage assembly
Sample holder plate
X-piezo stage
X-Y translation stage
Sample holder stage platform
Adapter plate
6x4 inches bread board
Sample holder stage assembly
Cube holder plate White light
QPD
Dichroic holder cube LC6W
LB6C plate
SM2D25Daperture
LCP02adapter
LB4C mirror holder
Condenser and objective
XZ
Y
QPD lens L5 STIXY translation stageQPD
QPD platform
Optmechanical controls
Base stage
Clamp
QPD sensor assembly
XZ
YQPD
Y
X
L5
STIXY translation stage
Optmechanical controls
LCP02
QPD platform
LFND
QPD sensor assembly