Alex Dooraghi

Micro-chemical synthesis of molecular probes on an electronic microfluidic device

We have developed an all-electronic digital microfluidic device for microscale chemical synthesis in organic solvents, operated by electrowetting-on-dielectric (EWOD). As an example of the principles, we demonstrate the multistep synthesis of [18F]FDG, the most common radiotracer for positron emission tomography (PET), with high and reliable radio-fluorination efficiency of [18F]FTAG (88 ± 7%, n = 11) and quantitative hydrolysis to [18F]FDG (> 95%, n = 11). We furthermore show that batches of purified [18F]FDG can successfully be used for PET imaging in mice and that they pass typical quality control requirements for human use (including radiochemical purity, residual solvents, Kryptofix, chemical purity, and pH). We report statistical repeatability of the radiosynthesis rather than best-case results, demonstrating the robustness of the EWOD microfluidic platform. Exhibiting high compatibility with organic solvents and the ability to carry out sophisticated actuation and sensing of reaction droplets, EWOD is a unique platform for performing diverse microscale chemical syntheses in small volumes, including multistep processes with intermediate solvent-exchange steps.

Optimization of microfluidic PET tracer synthesis with Cerenkov imaging

Microfluidic technologies provide an attractive platform for the synthesis of radiolabeled compounds. Visualization of radioisotopes on chip is critical for synthesis optimization and technological development. With Cerenkov imaging, beta particle emitting isotopes can be localized with a sensitive CCD camera. In order for Cerenkov imaging to also serve as a quantitative tool, it is necessary to understand how material properties relevant to Cerenkov emission, namely, index of refraction and beta particle stopping power, affect Cerenkov light output. In this report, we investigate the fundamental physical characteristics of Cerenkov photon yield at different stages of [(18)F]FDG synthesis on the electrowetting on dielectric (EWOD) microfluidic platform. We also demonstrate how Cerenkov imaging has enabled synthesis optimization. Geant4, a Monte Carlo program applied extensively in high energy physics, is used to simulate Cerenkov photon yield from (18)F beta particles traversing materials of interest during [(18)F]FDG synthesis on chip. Our simulations show that the majority (approximately two-thirds) of the (18)F beta particle energy available to produce Cerenkov photons is deposited on the glass plates of the EWOD chip. This result suggests the possibility of using a single calibration factor to convert Cerenkov signal to radioactivity, independent of droplet composition. We validate our simulations with a controlled measurement examining varying ratios of [(18)O]H2O, dimethyl sulfoxide (DMSO), and acetonitrile (MeCN), and find a consistent calibration independent of solvent composition. However, the calibration factor may underestimate the radioactivity in actual synthesis due to discoloration of the droplet during certain steps of probe synthesis. In addition to the attractive quantitative potential of Cerenkov imaging, this imaging strategy provides indispensable qualitative data to guide synthesis optimization. We are able to use this imaging technique to optimize the mixing protocol as well as identify and correct for loss of radioactivity due to the migration of radioactive vapor outside of the EWOD heater, enabling an overall increase in the crude radiochemical yield from 50 ± 3% (n = 3) to 72 ± 13% (n = 5).

Fast Metabolic Response to Drug Intervention Through Analysis on a Miniaturized, Highly Integrated Molecular Imaging System

We report on a radiopharmaceutical imaging platform designed to capture the kinetics of cellular responses to drugs. Methods: A portable in vitro molecular imaging system comprising a microchip and a β-particle imaging camera permitted routine cell-based radioassays of small numbers of either suspended or adherent cells. We investigated the kinetics of responses of model lymphoma and glioblastoma cancer cell lines to 18F-FDG uptake after drug exposure. Those responses were correlated with kinetic changes in the cell cycle or with changes in receptor tyrosine kinase signaling. Results: The platform enabled direct radioassays of multiple cell types and yielded results comparable to those from conventional approaches; however, the platform used smaller sample sizes, permitted a higher level of quantitation, and did not require cell lysis. Conclusion: The kinetic analysis enabled by the platform provided a rapid (∼1 h) drug screening assay.

A DOI Detector With Crystal Scatter Identification Capability for High Sensitivity and High Spatial Resolution PET Imaging

A new phoswich detector is being developed at the Crump Institute, aiming to provide improvements in sensitivity, and spatial resolution for PET. The detector configuration is comprised of two layers of pixelated scintillator crystal arrays, a glass lightguide and a light detector. The annihilation photon entrance (top) layer is a 48×48 array of 1.01 × 1.01 × 7 mm3 LYSO crystals. The bottom layer is a 32 × 32 array of 1.55 × 1.55 × 9 mm3 BGO crystals. A tapered, multiple-element glass lightguide is used to couple the exit end of the BGO crystal array (52 × 52 mm2) to the photosensitive area of the Position Sensitive Photomultiplier Tube (46 × 46 mm2), allowing the creation of flat panel detectors without gaps between the detector modules. Both simulations and measurements were performed to evaluate the characteristics and benefits of the proposed design. The GATE Monte Carlo simulation indicated that the total fraction of the cross layer crystal scatter (CLCS) events in singles detection mode for this detector geometry is 13.2%. The large majority of these CLCS events (10.1% out of 13.2%) deposit most of their energy in a scintillator layer other than the layer of first interaction. Identification of those CLCS events for rejection or correction may lead to improvements in data quality and imaging performance. Physical measurements with the prototype detector showed that the LYSO, BGO and CLCS events were successfully identified using the delayed charge integration (DCI) technique, with more than 95% of the LYSO and BGO crystal elements clearly resolved. The measured peak-to-valley ratios (PVR) in the flood histograms were 3.5 for LYSO and 2.0 for BGO. For LYSO, the energy resolution ranged from 9.7% to 37.0% full width at half maximum (FWHM), with a mean of 13.4 ± 4.8%. For BGO the energy resolution ranged from 16.0% to 33.9% FWHM, with a mean of 18.6 ± 3.2%. In conclusion, these results demonstrate that the proposed detector is feasible and can potentially lead to a high spatial resolution, high sensitivity and DOI PET system.

On-chip product purification for complete microfluidic radiotracer synthesis

Solid phase extraction was incorporated into an electrowetting-on-dielectric chip for radiochemical purification of a positron emission tomography tracer that was radiolabeled on the same chip. The radiotracer droplet was mixed with alumina particles, and the alumina particles were filtered out from the droplet through a line of pillars, all by electrowetting droplet movement. Fluorination reaction and on-chip purification were analyzed with both Cerenkov imaging and off-chip radio-thin layer chromatography measurements. The measurements were compared to test the validity of the combined use of filtration on-chip and Cerenkov imaging as an alternative approach for monitoring reaction yield without the need to extract sample from the chip.

Quantitative assessments of glycolysis from single cells

The most common positron emission tomography (PET) radio-labeled probe for molecular diagnostics in patient care and research is the glucose analog, 2-deoxy-2-[F-18]fluoro-D-glucose (18F-FDG). We report on an integrated microfluidics-chip/beta particle imaging system for in vitro18F-FDG radioassays of glycolysis with single cell resolution. We investigated the kinetic responses of single glioblastoma cancer cells to targeted inhibitors of receptor tyrosine kinase signaling. Further, we find a weak positive correlation between cell size and rate of glycolysis.

Evaluation of transimpedance amplifiers for readout of a position sensitive avalanche photodiode

BetaBox, a direct detection beta camera utilizing a 13.5×13.5mm2 active area position sensitive avalanche photodiode (PSAPD) (Radiation Monitoring Devices) is currently being developed to image distributions of charged particles in microfluidic chips. The previously reported signal processing chain incorporated charge sensitive amplifiers (Cremat Inc., CR-110) in the frontend readout design. Charge sensitive amplifiers, particularly the CR-110 product, are commonly used as preamplifiers when reading out PSAPDs. However, the 140µs pulse decay of this device suggests a possible count rate limitation in the performance of the beta camera. To address this limitation, the frontend readout was redesigned to replace the charge sensitive amplifiers with op-amps configured as transimpedance amplifiers. We hypothesize that, since the equivalent circuit of the PSAPD appears more like a charge division network loaded by a large semiconductor junction capacitance (∼150 pF) than an APD, this signal source can be optimally processed using the transimpedance amplifier approach. Results show that the designed signal processing chain using a transimpedance amplifier with 100ns RC time constant displays not only comparable image quality to the previous signal chain using the CR-110 but also reduced image distortion at high count rates.