Soft and Hard X-Ray Microscopy
David Attwood
University of California, Berkeley
andAdvanced Light Source, LBNL
Cheiron School
October 2010
SPring-8
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The short wavelength region of theelectromagnetic spectrum

n = 1 – δ + iβ

δ, β << 1
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Two common soft x-ray microscopes

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A Fresnel zone plate lens for x-ray microscopy

Erik Anderson, LBNL
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λ = 1.52 nm (815 eV)
Δr = 15 nm
N = 500
D = 30 µm
f = 300 µm
σ = 0.38
0.8 Δr = 12 nm

Cr/Si test pattern (Cr L3 @ 574 eV)(2000 X 2000,104 ph/pixel)
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Novel zone plates for specific functionality

Courtesy of Anne Sakdinawat, UC Berkeley
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High resolution zone plate microscopy• Well engineered• Sample indexing• Tiling for larger fieldof view
• Pre-focused• High sample throughput• Illumination important• Phase contrast

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The water window for biological
x-ray microscopy

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Fast freeze cryo fixation strongly mitigatesradiation dose effects

Helium passes through LN, is cooled,
and directed onto sample windows

Temperature (Celsius)

Fast Freeze
500

ΔT = – 50°c
16 ms
Δt-50
-100
-150
-100

10

20

30

4050

60

70

80

90

100

Time (milliseconds)

W. Meyer-Ilse, G. Denbeaux, L. Johnson, A. Pearson (CXRO-LBNL)
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Organelle details imaged with cryogenic
preservation and high spatial resolution
Cryo x-ray microscopy of 3T3 fibroblast cells
ERFilopodiaNucleus

Cell border
Nucleoli
C. Larabell, D. Yager, D. Hamamoto, M. Bissell, T. Shin (LBNL Life Sciences Division)
W. Meyer-Ilse, G. Denbeaux, L. Johnson, A. Pearson (CXRO-LBNL)
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Bending magnet radiation used with a soft x-ray microscope
to form a high resolution image of a whole, hydrated mouseepithelial cellhw = 520 eV
32 µm x 32 µm
Ag enhanced Au labelingof the microtubule network,color coded blue.
Cell nucleus and nucleoli,
moderately absorbing,coded orange.
Less absorbing aqueous
regions coded black.
W. Meyer-Ilse et al.
J. Microsc. 201, 395 (2001)
Courtesy of C. Larabell and W. Meyer-Ilse (LBNL)
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Bio-nanotomography for 3D imaging of cells
Nanotomography of
Cryogenic Fixed Cells

Soft X-Ray Nanotomographyof a Yeast Cell

Courtesy of G. Schneider (BESSY)
Surf. Rev. Lett. 9, 177 (2002)

λ = 2.4 nm
Courtesy of C. Larabell (UCSF & LBNL)
and M. LeGros (LBNL)
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Bio-nanotomography for 3D imaging of cells
Nanotomography ofCryogenic Fixed Cells

λ = 2.4 nm (517 eV)
Δr = 35 nm
N = 320NA = 0.034
D = 45 µm
f = 650 µm
σ = 0.64
Resolution = 60 nm

Soft X-Ray Nanotomographyof a Yeast Cell

λ = 2.4 nm
Courtesy of C. Larabell (UCSF & LBNL)
and M. LeGros (LBNL)
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Nanoscale 3-D biotomography
Mother daughter yeast cells just before separation
2-D slice from 3-D
Tomogram. Images
every 2°, 180° dataset, several minutes.
Δr = 45 nm
Color coding
identifies subcellularcomponents by their
x-ray absorption
coefficients
Courtesy of Carolyn Larabell, UCSF/LBNL.
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Applications of soft x-ray microscopy
Biotomography at 60 nm resolution• Cryofixation• 2° angular intervals• Depth of focus limits resolution• New XM-2 dedicated to biologicalapplications, will become major
facility worldwide to draw biologists
to this evolving capability

Courtesy of C. Larabell (UCSF & LBNL)

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Magnetic x-ray microscopy using x-ray
magnetic circular dichroism (XMCD)

Magnetic X-Ray Microscopy• High spatial resolution in transmission• Bulk sensitive (thin films)• Complements surface sensitive PEEM
• Good elemental sensitivity• Good spin-orbit sensitivity

100 nmlines &spaces• Allows applied magnetic field• Insensitive to capping layers• In-plane and out-of-plane measurements

1 µm

Courtesy of P. Fischer, (MPI, Stuttgart) and G. Denbeaux (CXRO/LBNL)
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Magnetic domains imaged at different
photon energies
FeGd MultilayerContrast
reversal

1 µmħω = 704 eV
below Fe L-edgesħω = 707.5
eVFe L3-edgeħω = 720.5
eVFe L2-edge

P. Fischer, T. Eimueller, M. Koehler (U. Wuerzberg)
S. Tsunashima (U. Nagoya) and N. Tagaki (Sanyo)
G. Denbeaux, L. Johnson, A. Pearson (CXRO-LBNL)
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Magnetic recording of nanomagnetic patterns
to 15 nm spatial resolution
CoCrPt alloy
Co L3-edge at 778 eV(1.59 nm)

200 nmCourtesy of Peter Fischer (LBNL)
P. Fischer et al., Mat. Today 9, 26 (2006).

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Time resolved studies of vortex dynamics inpatterned permalloy thin films
Pump and Probe setup requires:• Pump: Current pulse to “pump” sample• Probe: X-ray pulses (70ps) from ALS 2
Bunch mode
• Perfect repeatability of dynamics

Sample:50 nm thick 2µmx4µm
permalloy (Ni80Fe20)
100nm thick gold waveguide(∆I along waveguide generates field to pumpsample)

BeforePump(t=0)

t=+1 ns

B.L. Mesler, P. Fischer,W. Chao, E. H. Anderson,
D.H. Kim J. Vac. Sci. Technol. B 25, 2598 (2007).

t=+1.6 ns

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Environmental Consequencesof Portland cement
1.5 billion ton of cement

Generates 1.5 billion
ton of CO2
Responsible for 7%
CO2 production inthe world

Courtesy of Professor Paulo Monteiro, CEE, UC Berkeley

Problem!

Nanoscale x-ray imaging of cement processes:
early hydrates forming during the pre-induction period
1 µm

Early hydrates(Sheaf of wheat)Grain

C3S hydrated for 34 min. insaturated lime and calcium
sulfate at w/c = 5, 1 s
exposure time, 516 eV, scalebar 1µm.
Courtesy of Professor Paulo Monteiro, CEE, UC Berkeley
EE298 Seminar/ David Attwood / April 17, 2010

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Nanoscale x-ray imaging of cement processes

13 min34 min50 min10 min20 min1 hr 49 min

10 min34 min1 hr 8 min

Courtesy of Professor Paulo Monteiro, CEE, UC Berkeley
EE298 Seminar/ David Attwood / April 17, 2010

520 eV, 40 nm – spatial resolution
EE298_Seminar_17Apr2010.ppt

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Spectromicroscopy: high spatial and highspectral resolution of surface and thin films

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Biofilm from Saskatoon RiverRESULTS• Ni, Fe, Mn, Ca, K, O, C elemental map,
(there was no sign of Cr.)• Different oxidation states for Fe and Ni

Fe 2pOD
1.5

1.0

1 µm

0.55 µm

Protein (gray), Ca, K

0700

705710

715

720

725

Different oxidation states (minerals) found for Fe & Ni

Tohru Araki, Adam Hitchcock (McMaster University)
Tolek Tyliszczak, LBNL
Sample from: John Lawrence, George Swerhone (NWRISaskatoon), Gary Leppard (NWRI-CCIW)
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Patterned polymer photoresists
ALS-MES 11.0.2

M.K. Gilles, R. Planques, S.R. Leone
LBNL
Samples from B. Hinsberg, F. Huele
IBM Almaden

280 eV

290 eV

Exposure to UV light results in loss of carbonyl peak

Map chemical spectra taken of pure samples
onto a sample containing both components
Courtesy of Mary Gilles, LBNL
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Hard x-ray zone plate microscopy• Shorter wavelengths, potentially better spatial
resolution and greater depth-of-field.
• Less absorption (β); phase shift (δ) dominates,higher efficiency.
• Thicker structures required (e.g., zones), higheraspect ratios pose nanofabrication challenges.
• Contrast of nanoscale samples minimal; will require
good statistics, uniform background, dose mitigation.

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nanoXCT: Schematic and Challenges
X-ray Zone-plate Lens

Challenges for achieving nm scale resolution:• High resolution objective lens: limiting the ultimate resolution• High numerical aperture condenser lens:• Detector: high efficiency for lab. source and high speed for synchrotron sources• Precision mechanical system
Courtesy of Wenbing Yun and Michael Feser, Xradia

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Xradia nanoXCT: Sub-25 nm Hard X-ray Image
Xradia Resolution Pattern• 50 nm bar width• 150 nm thick Au• 8keV x-ray energy• 3rd diffraction order

F. Duewer, M. Tang,
G. C. Yin, W. Yun,
M. Feser, et al.

Xradia nano-XCT
8-50S installed atNSRRC, Taiwan

400 nm2

Elemental contrast by tuning energy across thecopper absorption edge ( Guan-Chian Yin et al)

8.4 keV

9.5 keV

9.5

8.4

Intensity difference between
E = 8.4 keV and 9.5 keV
3Tomography of a Tungsten plug with “keyhole” at ~60 nm spatial resolution

Xradia and NSRRC

APL. 88, 241115 (2006)

Scanning x-ray fluorescence microprobe (µ-XRF)

J.H. Underwood and A.C. Thompson, NIM A266, 296 & 318 (1988).
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X-ray microprobe at SPring-8
Optical microscopemm

f1: 252

PIN photodiodemm
f2: 150

98m
45m

SDD
Mirror manipulator

DCMUndulator

TC1SlitIon chamber
Incident Slit

Beam monitor
Sample & Scanner

Experimental hutch

S. Matsuyama et al.,Rev. Sci. Instrum.
77, 103102 (2006)

Front end

Courtesy of K. Yamauchi andH. Mimura, Osaka University.
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Sub-cellular elemental analysis using the hard
x-ray fluorescence microprobe at SPring-8

S. Matsuyama et al.,
X-Ray Spectrom.(2010).

Courtesy of K. Yamauchi andH. Mimura, Osaka University.
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8Breaking the 10 nm barrier in hard x-ray focusing
In-situ phase compensation

7 nm

Piezo-electric phasecompensator

Focusing mirror with phase error

Focusing mirror with phase error
H. Mimura et al., Nature Physics, 6, 122 (2009)
XRM2010, 16th, Aug, 2010

Optical configuration for active phase compensation
Knife edge scanner

Fizeau interferometer

9APD@BL29XUL

X-ray

Deformable mirror

Focusing mirror(Mirror B)

Opticalinterferometer(ZYGO GPI)

CCD camera(for alignment)

Focusing mirror
Focalpoint

Ion chamber

Adaptive mirror

150mm

Zooming tube+ X-ray CCD

Zooming tube + CCD

APD placed at dark field

250mm

H. Mimura et al., Nature Physics, 6, 122 (2009)

XRM2010, 16th, Aug, 2010

Synchrotron-based art conservation at ESRFµ-XRF, µ-XRD, µ-XANES

Courtesy of Marine Cotte (ESRF, Grenoble, France)
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Examples of µ-XANES K-edge spectra
occurring in art materials
SulfurChlorine

Courtesy of Marine Cotte (ESRF, Grenoble, France)
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18th Dynasty Egyptian glass vase studied for anunderstanding of color and opaqueness in antiquity
1st production of glass objects Egypt (1500 B.C.),
opaque, colored, nanoscale calcium antimonate

Courtesy of Marine Cotte (ESRF, Grenoble, France)
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Synchrotron radiation x-ray tomographic microscopy
Swiss Light Source (SRXTM)

Courtesy of Marco Stampanoni, Swiss Light Source.

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TOMCAT Microscope
Swiss Light Source

1 micron @ 10% MTF reached routinely
Courtesy of Marco Stampanoni, Swiss Light Source.

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TOMCAT Microscope
Swiss Light Source

To CCD…

scintillator

objective

mirror

X-ray beam

rotation stage

centering device

1 micron @ 10% MTF reached routinely
Courtesy of Marco Stampanoni, Swiss Light Source.

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Hard x-ray 3D x-ray tomography: microvascular
Swiss Light Source architecture of a mouse brain

SRXTM, 25 keV,
15 µm resolution
Tomcat Beamline,
Swiss Light Source
M. Stampanoni,T. Krucker et al.,Adv. Neur. Res. (2008)

Courtesy of Marco Stampanoni, Swiss Light Source.

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Tomographic reconstruction of a 500 million year
Swiss Light Source old fossilized embryo from Southern China
Markelia hunanensis
relative of modern roundworms
and arthropods

SRXTM, 17.5 keV,
15 µm resolution
Tomcat Beamline,
Swiss Light Source
P. Donoghue, S. Bengtson, M.
Stampanoni et al., Nature 442, (2006)
Courtesy of Marco Stampanoni, Swiss Light Source.

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A lens is not necessarily requiredΔrresol. = k1 λ / NA

“Lensless” coherent diffraction imaging (CDI) is being
aggressively pursued.

Frontiers in Optics, OSA, San Jose, CA, October 15, 20093

Coherent diffractive imaging (CDI)

Frontiers in Optics, OSA, San Jose, CA, October 15, 20094

Coherent diffractive imaging (CDI) examples
Femtosecond diffractive imaging
with a free electron laser

CDI with laboratory scalehigh harmonic generation (HHG)

Flash FEL, λ = 32 nm (39 eV)
25 fsec, 1012 photons/pulse
62 nm resolution

HHG, n = 27, λ= 29 nm (43 eV )94 nm spatial resolution

Chapman, et al. Nature Physics (2006)

Sandberg, et al. PNAS (2008)

Synchrotron based CDI of 100 nm Au spheres

Synchrotron CDI of Au particles
λ = 0.083 nm (15 keV),
5 nm “resolution”
Schroer, et al. PRL (2008)

Synchrotron based CDI of a freeze dried yeast cell
CDI
ALS/9.0.1
λ = 1.66 nm(750 eV)

STXM
NSLS/X1
λ = 2.3 nm(540 eV)
42 nm
resolution

Tilted 3°

Tilted 4°

Shapiro, et al. PRL (2005)

30 nm fine features

Frontiers in Optics, OSA, San Jose, CA, October 15, 20095

Lensless imaging of magnetic nanostructuresby x-ray spectro-holography

Frontiers in Optics, OSA, San Jose, CA, October 15, 20096

Lectures online at www.youtube.com

Amazon.com

UC Berkeley
www.coe.berkeley.edu/AST/sxreuv
www.coe.berkeley.edu/AST/srms
www.coe.berkeley.edu/AST/sxr2009

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