I i nstrumentat on for ESR M icroscopy Microscopy CURT R DUNNAM 1

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1 Instrumentation for ESR Microscopy CURT R DUNNAM 1

2 ACERT ESRM Instrumentation Development elopment to Date: CW (9 GHz) Modified Varian E-series X-band Spectrometer (add-on system) stem) Capable of Projection Reconstruction (PR) and Modulated Field Gradient (MFG), spectral- spatial imaging modalities Resolution, LiPc phantom: ~10μm, 64x64 pixels, image acquisition iti times of f6 min (PR) and 34 min (MFG) (2004) 2

3 ACERT CW ESRM, cont d: 3

4 ACERT ESRM Instrumentation Development elopment to Date, cont d: PULSED (16 GHz) R.F., Timing, Gradient Driver systems and system control software designed specifically for pulsed ESRM Peak gradient capability Gx ~75 T/m, Gy ~47 T/m Sub-micron resolution, ~0.7x0.75x7.5μm obtained for high-spin sample with image size of 250x250x64 voxels Resolution ~3x3x10μm μ for ~1mM tritylox063 _ d24 solution; ~10x10x50 μm for ~0.5mM 15 N PDT Imaging of cancerous tissue and cell suspensions has been demonstrated 4

5 PULSED ESR Microscope Instrument: Pi Principal i lsystems: BRIDGE GRADIENT DRIVER SYSTEM HV PREREGULATOR HAVERSINE DRIVER PROBE DIGITIZER/AVERAGER 5

6 ACERT PULSED ESRM, cont d: 6

7 ESRM INSTRUMENTATION DEVELOPMENT at ACERT DRIVEN BY BIOMEDICAL AND CLINICAL APPLICATIONS, e.g., Controlled drug release (Blank, et al., 2005) Cancerous tissue imaging (Oxygenation) 02 and redox metabolism Assessment of radiation damage Cellular l imaging i (with contrast t agent) O2 - permeable biomaterials investigation 7

8 ESRM DEVELOPMENT AIMS Advance pulsed ESRM to true micron resolution level (1 x 1 x 3 µm) for 2-D images of biologically-relevant samples. This will be done in several independent steps, all designed to improve the SNR: Increase the pulse sequence repetition rate Generate shorter, more intense pulse gradients Improved probe and resonator designs Construction of a Ka-Band ESR microscope Evaluate new pulse modalities using new pulse gradient capabilities Develop new sample preparation p methods for biomedical and clinical applications Conduct ESR microscopy of tissue and cell samples with trityl and nitroxide probe derivatives 8

9 ESRM Instrument Development Step: Increased Repetition Rate Pulse sequence repetition rate is currently limited to KHz by gradient driver and probe dissipation considerations. Repetition rate increase by a factor of 4-40 x, to a maximum rate of KHz is possible: Conduct heat away from critical probe elements by means of heat-conductive encasing material, heat pipes, etc. Provide ample cooling ggas or air flow through the resonator assembly. Replace high tempco dielectric ring resonator with alternate type, eg e.g., loop gap, planar. 9

10 ESRM Instrument Development Step: Shorter, more intense pulse gradients Pulsed haversine gradients Gx, Gy are currently limited to ca 75 T/m, 47 T/m, respectively, min. duration 1us. Frequency encoding ggradient Gz is currently limited to ca 10 T/m. Near term: increase Gx, Gy intensity by ca 1.5 while decreasing period proportionately. Future: Add high intensity pulsed haversine drive capability for Gz Increase haversine drive capability to ca 2x current max. Implement hybrid driver for fast rectangular gradient scheme 10

11 Hybrid Gradient Driver Fast rectangular gradient simulation: Tr, Tf ~ 80ns 11

12 ESRM Instrument Development Step: Improved probe and resonator design Probe: Thermal management features Front-end LNA Resonator: Single Dielectric i Ring (superior filling factor) Loop Gap, planar (superior filling factor; low temperature coefficient; design flexibility, e.g.,fine-needle needle compatibility 12

13 Resonator Candidate Structures LGR at 15GHz z r= 750um w=650um z=1000um d t=100um w d=300um (distance from LGR to r coupling loop) t L~1.326nH, C~ pf - expected resonant frequency ~16.17GHz17GHz R~0.17Ohm Critically coupled loaded Excitation ti port with Q~400 impedance of 50Ohms 13

14 Resonator Candidate Structures, cont d Modified loop gap resonator w/ microfluidic channels E H C. Shin, 2008 H-field or E-field coupling is possible. (drawing is for E-field coupling) Resonator Sample chamber Gas permeable teflon tape Stripline Sample preparation would be similar to what we have now. This can be done for high frequency where glass capillary can not be used. 14

15 ESRM Instrument Development Step: Construction of a CW/Pulsed Ka-Band Microscope We plan to construct an integrated CW/pulsed ESR microscopy system operating at ca 35 GHz, primarily to take advantage of ω ω 03 scaling in SNR: SNR pulse 2μ V ηω M Q 8 0 c 0 k T Δ f b ω L 0 15

16 CW/PULSED Ka-Band Microscope: 16

17 ESR DEVELOPMENT AIMS, Cont d: Evaluation of new pulse imaging modalities to improve performance with shorter, more intense gradient pulses: SPI (after Subramanian, et al., 2002) Phase-encoding time is held constant for the image data collection, so the spectral information is inherently deconvolved, providing well-resolved pure spatial images that are relatively free of artifacts. Multiple-gradient mode (increment, delay time) may be employed to derive pixel-wise T2* information Multiple-point echo-based SPI for T2 evaluation and po2 analysis by comparison with intrinsic T2 FID For spin probes with T2* close to T2, ESRM based on FID collection can give an additional improvement factor of 2 in the signal amplitude. 17

18 ESRM Instrumentation Summary State-of-the-art ESRM performance for high-spinh i samples is currently atthe levell of sub-micron resolution. ESR microscope performance approaching the micron level for biologically-relevant samples appears to be a feasible objective in the context of a well-defined development effort based on existing ESRM instrumentation designs. 18

19 Acknowledgements: With special appreciation for their ongoing patient and insightful i ldiscussion i of fesr imaging and microscopy theory and application: Aharon Blank, Technion Peter Borbat, ACERT Chang Shin, LBL Sankaran Subramanian, NCI NIH/NCRR Grant P41RR