AFM Images
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Transcript of AFM Images
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AFM Images
Bacteria
Mosquito eye
DNA molecules
http://www.afmhelp.com/index.php?option=com_content&view=article&id=51&Itemid=57
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Today’s Topic: AFM(Next Time: FIONA)
• Experimental Approach via Atomic Force Microscopy• AFM: Can see Nanometer & Angstrom scale changes!• How you can see this: Hook’s Law• Imaging Mode, Force Mode.• To minimize noise, go to a higher frequency :
noise always goes like 1/f (f = frequency)• Force Mode: Worm Like Chain model of Protein Folding (and
DNA)
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http://cp.literature.agilent.com/litweb/pdf/5990-3293EN.pdf
AFM—Imaging and/or Force
http://www.home.agilent.com/agilent/editorial.jspx?cc=US&lc=eng&ckey=1774141&nid=-33986.0.02&id=1774141
Ionic repulsion,bends tip
Imaging – ScanMuller,
Biochemistry, 2008
Measure (z-axis) distance,Or
Constant force (by altering distance with feedback)
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http://cp.literature.agilent.com/litweb/pdf/5990-3293EN.pdf
AFM—Imaging and/or Force
http://www.home.agilent.com/agilent/editorial.jspx?cc=US&lc=eng&ckey=1774141&nid=-33986.0.02&id=1774141
Force – one place
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http://www.home.agilent.com/agilent/editorial.jspx?cc=US&lc=eng&ckey=1774141&nid=-33986.0.02&id=1774141
*
* Regular resolution; Super-resolution, ~ 10 nm at somewhat greater expense (>$150k)
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Diff. types of Microscopes(Diff. , techniques, diff. resolution)
MBC, Fig. 4-
1000x, 0.2 um 106x, 2 nm 20,000x 10 nm (3-D)
Or with CCD
TransmittedLight
Resolution of microscope ~/2, where is different for different techniques
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Measuring forcesThe force is not measured directly, but calculated by measuring the deflection of the lever, and knowing the stiffness of the cantilever. Hook’s law gives F = -kz, where F is the force, k is the stiffness of the lever, (in Newtons/meter) and z is the distance the lever is bent.
http://cp.literature.agilent.com/litweb/pdf/5990-3293EN.pdf
Most AFM probes are made from silicon and/or silicon nitride (Si3N4) wafers using semiconductor-based etching processes.
Hook’s Law and AFM
http://www.home.agilent.com/agilent/editorial.jspx?cc=US&lc=eng&ckey=1774141&nid=-33986.0.02&id=1774141
Ionic repulsion,bends tip
Imaging—ScanForce– one place
Can measure Angstrom resolution – Nobel Prize, 1986
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What determines how accurately you can measure?
If photodetector was on the moon, could you see infinitely small changes?
Brownian Noise & Equipartition Theorem
Each degree of freedom in Energy goes as x2 or v2 : has ½ kBT of energy.
A gas molecule at temperature T: Kinetic Energy: ½ mv2 : Has ½ kBT of energy
½ mv2 = ½ kT : v2 = kBT/m v = √kT/m : ~ 1000 mph!
If you have a spring at finite temperature:
Examples?
½ kx2 = ½ kBT
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AFM CantileverHow small of a motion can you measure?
Bend a cantilever (in z-direction):½ kz2 = ½ kBT
(z2 is the mean square deflectionof the cantilever caused by thermal vibrations)
z2 = kBT/k = 0.64Å/√k at 22˚C (where k is in N/m)
Say, 1 N/m = 1 nN/nm: 1 nN causes deviation of 1 nm
1 nN really large for bio0.01 N/m = 10 pN/nm: 1 pN causes a deviation of 1 nm)
Can measure an Angstrom or less!!
k between 0.001 to 100 N/m (Huge range! Very useful for measuring large range ∆z: F from 1 pN - nN)
z
k= 0.25Ewh3/L3, where E = modulus of Elasticity (Non-trivial but based on F= kL) --see Joe Howard,
Mechanics of Motor Proteins…
What is k?
(E, how stiff the material is).
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ΔX ~ (In1-In2) / (In1 + In2)
ΔY ~ (Out1-Out2) /(Out1+Out2)
N
P
N
P
In1 In2
Out1
Out2
POSITION
SIGNAL
Position sensitive detector (PSD)
Over a fairly wide range, it’s linear
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Typically, probe radius varies from 5 to 20 nm
http://webserv.jcu.edu/chemistry/faculty/waner/research/AFM/tipconv.htm
If tip size is large, have to worry about distortions
Convolution
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Convolution of tip and sample size
http://webserv.jcu.edu/chemistry/faculty/waner/research/AFM/tipconv.htm
Tobacco Mosaic Virus (TMV)
In truth, diameter of 180 Å. Due to finite tip size, w~ 350 A
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Cross-correlation
Cross-correlation
Correlation functions
http://www.scholarpedia.org/article/1/f_noise
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The End