Benefits and challenges of in vivo EPR nail biodosimetry in a second tier of medical triage in response to a large radiation event.

Following large-scale radiation events, an overwhelming number of people will potentially need mitigators or treatment for radiation-induced injuries. This necessitates having methods to triage people based on their dose and its likely distribution, so life-saving treatment is directed only to people who can benefit from such care. Using estimates of victims following an improvised nuclear device striking a major city, we illustrate a two-tier approach to triage. At the second tier, after first removing most who would not benefit from care, biodosimetry should provide accurate dose estimates and determine whether the dose was heterogeneous. We illustrate the value of using in vivo electron paramagnetic resonance nail biodosimetry to rapidly assess dose and determine its heterogeneity using independent measurements of nails from the hands and feet. Having previously established its feasibility, we review the benefits and challenges of potential improvements of this method that would make it particularly suitable for tier 2 triage. Improvements, guided by a user-centered approach to design and development, include expanding its capability to make simultaneous, independent measurements and improving its precision and universality.

Developments in Biodosimetry Methods for Triage With a Focus on X-band Electron Paramagnetic Resonance In Vivo Fingernail Dosimetry.

Instrumentation and application methodologies for rapidly and accurately estimating individual ionizing radiation dose are needed for on-site triage in a radiological/nuclear event. One such methodology is an in vivo X-band, electron paramagnetic resonance, physically based dosimetry method to directly measure the radiation-induced signal in fingernails. The primary components under development are key instrument features, such as resonators with unique geometries that allow for large sampling volumes but limit radiation-induced signal measurements to the nail plate, and methodological approaches for addressing interfering signals in the nail and for calibrating dose from radiation-induced signal measurements. One resonator development highlighted here is a surface resonator array designed to reduce signal detection losses due to the soft tissues underlying the nail plate. Several surface resonator array geometries, along with ergonomic features to stabilize fingernail placement, have been tested in tissue-equivalent nail models and in vivo nail measurements of healthy volunteers using simulated radiation-induced signals in their fingernails. These studies demonstrated radiation-induced signal detection sensitivities and quantitation limits approaching the clinically relevant range of ≤ 10 Gy. Studies of the capabilities of the current instrument suggest that a reduction in the variability in radiation-induced signal measurements can be obtained with refinements to the surface resonator array and ergonomic features of the human interface to the instrument. Additional studies are required before the quantitative limits of the assay can be determined for triage decisions in a field application of dosimetry. These include expanded in vivo nail studies and associated ex vivo nail studies to provide informed approaches to accommodate for a potential interfering native signal in the nails when calculating the radiation-induced signal from the nail plate spectral measurements and to provide a method for calibrating dose estimates from the radiation-induced signal measurements based on quantifying experiments in patients undergoing total-body irradiation or total-skin electron therapy.

Dielectric-Backed Aperture Resonators for X-Band in vivo EPR Nail Dosimetry.

A new resonator for X-band in vivo EPR nail dosimetry, the dielectric-backed aperture resonator (DAR), is developed based on rectangular TE102 geometry. This novel geometry for surface spectroscopy improves at least a factor of 20 compared to a traditional non-backed aperture resonator. Such an increase in EPR sensitivity is achieved by using a non-resonant dielectric slab, placed on the aperture inside the cavity. The dielectric slab provides an increased magnetic field at the aperture and sample, while minimizing sensitive aperture resonance conditions. This work also introduces a DAR semi-spherical (SS)-TE011 geometry. The SS-TE011 geometry is attractive due to having twice the incident magnetic field at the aperture for a fixed input power. It has been shown that DAR provides sufficient sensitivity to make biologically relevant measurements both in vitro and in vivo Although in vivo tests have shown some effects of physiological motions that suggest the necessity of a more robust finger holder, equivalent dosimetry sensitivity of approximately 1.4 Gy has been demonstrated.

Development and validation of an ex vivo electron paramagnetic resonance fingernail biodosimetric method.

There is an imperative need to develop methods that can rapidly and accurately determine individual exposure to radiation for screening (triage) populations and guiding medical treatment in an emergency response to a large-scale radiological/nuclear event. To this end, a number of methods that rely on dose-dependent chemical and/or physical alterations in biomaterials or biological responses are in various stages of development. One such method, ex vivo electron paramagnetic resonance (EPR) nail dosimetry using human nail clippings, is a physical biodosimetry technique that takes advantage of a stable radiation-induced signal (RIS) in the keratin matrix of fingernails and toenails. This dosimetry method has the advantages of ubiquitous availability of the dosimetric material, easy and non-invasive sampling, and the potential for immediate and rapid dose assessment. The major challenge for ex vivo EPR nail dosimetry is the overlap of mechanically induced signals and the RIS. The difficulties of analysing the mixed EPR spectra of a clipped irradiated nail were addressed in the work described here. The following key factors lead to successful spectral analysis and dose assessment in ex vivo EPR nail dosimetry: (1) obtaining a thorough understanding of the chemical nature, the decay behaviour, and the microwave power dependence of the EPR signals, as well as the influence of variation in temperature, humidity, water content, and O2 level; (2) control of the variability among individual samples to achieve consistent shape and kinetics of the EPR spectra; (3) use of correlations between the multiple spectral components; and (4) use of optimised modelling and fitting of the EPR spectra to improve the accuracy and precision of the dose estimates derived from the nail spectra. In the work described here, two large clipped nail datasets were used to test the procedures and the spectral fitting model of the results obtained with it. A 15-donor nail set with 90 nail samples from 15 donors was used to validate the sample handling and spectral analysis methods that have been developed but without the interference of a native background signal. Good consistency has been obtained between the actual RIS and the estimated RIS computed from spectral analysis. In addition to the success in RIS estimation, a linear dose response has also been achieved for all individuals in this study, where the radiation dose ranges from 0 to 6 Gy. A second 16-donor nail set with 96 nail samples was used to test the spectral fitting model where the background signal was included during the fitting of the clipped nail spectra data. Although the dose response for the estimated and actual RIS calculated in both donor nail sets was similar, there was an increased variability in the RIS values that was likely due to the variability in the background signal between donors. Although the current methods of sample handling and spectral analysis show good potential for estimating the RIS in the EPR spectra of nail clippings, there is a remaining degree of variability in the RIS estimate that needs to be addressed; this should be achieved by identifying and accounting for demographic sources of variability in the background nail signal and the composition of the nail matrix.

Electron paramagnetic resonance dosimetry for a large-scale radiation incident.

With possibilities for radiation terrorism and intensified concerns about nuclear accidents since the recent Fukushima Daiichi event, the potential exposure of large numbers of individuals to radiation that could lead to acute clinical effects has become a major concern. For the medical community to cope with such an event and avoid overwhelming the medical care system, it is essential to identify not only individuals who have received clinically significant exposures and need medical intervention but also those who do not need treatment. The ability of electron paramagnetic resonance to measure radiation-induced paramagnetic species, which persist in certain tissues (e.g., teeth, fingernails, toenails, bone, and hair), has led to this technique becoming a prominent method for screening significantly exposed individuals. Although the technical requirements needed to develop this method for effective application in a radiation event are daunting, remarkable progress has been made. In collaboration with General Electric and through funding committed by the Biomedical Advanced Research and Development Authority, electron paramagnetic resonance tooth dosimetry of the upper incisors is being developed to become a Food and Drug Administration-approved and manufacturable device designed to carry out triage for a threshold dose of 2 Gy. Significant progress has also been made in the development of electron paramagnetic resonance nail dosimetry based on measurements of nails in situ under point-of-care conditions, and in the near future this may become a second field-ready technique. Based on recent progress in measurements of nail clippings, it is anticipated that this technique may be implementable at remotely located laboratories to provide additional information when the measurements of dose on-site need to be supplemented. The authors conclude that electron paramagnetic resonance dosimetry is likely to be a useful part of triage for a large-scale radiation incident.

Advances towards using finger/toenail dosimetry to triage a large population after potential exposure to ionizing radiation.

Rapid and accurate retrospective dosimetry is of critical importance and strategic value for the emergency medical response to a large-scale radiological/nuclear event. One technique that has the potential for rapid and accurate dosimetry measurements is electron paramagnetic resonance (EPR) spectroscopy of relatively stable radiation-induced signals (RIS) in fingernails and toenails. Two approaches are being developed for EPR nail dosimetry. In the approach using ex vivo measurements on nail clippings, accurate estimation of the dose-dependent amplitude of the RIS is complicated by the presence of mechanically-induced signals (MIS) that are generated during the nail clipping. Recent developments in ex vivo nail dosimetry, including a thorough characterization of the MIS and an appreciation of the role of hydration and the development of effective analytic techniques, have led to improvements in the accuracy and precision of this approach. An in vivo nail dosimetry approach is also very promising, as it eliminates the problems of MIS from the clipping and it has the potential to be an effective and efficient approach for field deployment. Two types of EPR resonators are being developed for in vivo measurements of fingernails and toenails.

A Deployable In Vivo EPR Tooth Dosimeter for Triage After a Radiation Event Involving Large Populations.

In order to meet the potential need for emergency large-scale retrospective radiation biodosimetry following an accident or attack, we have developed instrumentation and methodology for in vivo electron paramagnetic resonance spectroscopy to quantify concentrations of radiation-induced radicals within intact teeth. This technique has several very desirable characteristics for triage, including independence from confounding biologic factors, a non-invasive measurement procedure, the capability to make measurements at any time after the event, suitability for use by non-expert operators at the site of an event, and the ability to provide immediate estimates of individual doses. Throughout development there has been a particular focus on the need for a deployable system, including instrumental requirements for transport and field use, the need for high throughput, and use by minimally trained operators.Numerous measurements have been performed using this system in clinical and other non-laboratory settings, including in vivo measurements with unexposed populations as well as patients undergoing radiation therapies. The collection and analyses of sets of three serially-acquired spectra with independent placements of the resonator, in a data collection process lasting approximately five minutes, provides dose estimates with standard errors of prediction of approximately 1 Gy. As an example, measurements were performed on incisor teeth of subjects who had either received no irradiation or 2 Gy total body irradiation for prior bone marrow transplantation; this exercise provided a direct and challenging test of our capability to identify subjects who would be in need of acute medical care.

Cerebral oxygenation in awake rats during acclimation and deacclimation to hypoxia: an in vivo electron paramagnetic resonance study.

Exposure to high altitude or hypobaric hypoxia results in a series of metabolic, physiologic, and genetic changes that serve to acclimate the brain to hypoxia. Tissue Po(2) (Pto(2)) is a sensitive index of the balance between oxygen delivery and utilization and can be considered to represent the summation of such factors as cerebral blood flow, capillary density, hematocrit, arterial Po(2), and metabolic rate. As such, it can be used as a marker of the extent of acclimation. We developed a method using electron paramagnetic resonance (EPR) to measure Pto(2) in unanesthetized subjects with a chronically implanted sensor. EPR was used to measure rat cortical tissue Pto(2) in awake rats during acute hypoxia and over a time course of acclimation and deacclimation to hypobaric hypoxia. This was done to simulate the effects on brain Pto(2) of traveling to altitude for a limited period. Acute reduction of inspired O(2) to 10% caused a decline from 26.7 ± 2.2 to 13.0 ± 1.5 mmHg (mean ± SD). Addition of 10% CO(2) to animals breathing 10% O(2) returned Pto(2) to values measured while breathing 21% O(2,) indicating that hypercapnia can reverse the effects of acute hypoxia. Pto(2) in animals acclimated to 10% O(2) was similar to that measured preacclimation when breathing 21% O(2). Using a novel, individualized statistical model, it was shown that the T(1/2) of the Pto(2) response during exposure to chronic hypoxia was approximately 2 days. This indicates a capacity for rapid adaptation to hypoxia. When subjects were returned to normoxia, there was a transient hyperoxygenation, followed by a return to lower values with a T(1/2) of deacclimation of 1.5 to 3 days. These data indicate that exposure to hypoxia results in significant improvements in steady-state oxygenation for a given inspired O(2) and that both acclimation and deacclimation can occur within days.

Surface loop resonator design for in vivo EPR tooth dosimetry using finite element analysis.

Finite element analysis is used to evaluate and design L-band surface loop resonators for in vivo electron paramagnetic resonance (EPR) tooth dosimetry. This approach appears to be practical and useful for the systematic examination and evaluation of resonator configurations to enhance the precision of dose estimates. The effects of loop positioning in the mouth are examined, and it is shown that the sensitivity to loop position along a row of molars is decreased as the loop is moved away from the teeth.

Oxygen effects on the EPR signals from wood charcoals: experimental results and the development of a model.

Charcoals prepared from certain tropical woods contain stable paramagnetic centers, and these have been characterized by EPR spectroscopy in the absence and presence of oxygen. The EPR-detectable spin density has been determined, as has been the temperature- and frequency-dependence of the oxygen broadening of the EPR signal, which is orders of magnitude larger than that observed with other materials, such as lithium phthalocyanine. Three Lorentzian components are required to fit the char EPR spectrum in the presence of oxygen, and the oxygen-dependence of the line width, intensity, and resonance position of the three components have been quantified. These results and the properties of porous carbonaceous materials are used to develop a model to explain the effect of oxygen on the char EPR spectral properties. The model is based on oxygen adsorption on the char surface according to a Langmuir isotherm and a dipolar interaction between the paramagnetic adsorbed gas and the charcoal spins. The three EPR components are correlated with the three known classes (sizes) of pores in charcoal, with the largest line broadening attributed to dipolar relaxation of spins in micropores, which have a larger specific surface area and a higher concentration of adsorbed oxygen. An attenuated, but similar, EPR response to oxygen by chars when they are immersed in aqueous solution is attributed to water competition with oxygen for adsorption on the char surface.

The effect of oxygen therapy on brain damage and cerebral pO(2) in transient focal cerebral ischemia in the rat.

We examined the effect of hyperbaric oxygen (HBO) and normobaric oxygen (NBO) on neurologic damage and brain oxygenation before and after focal cerebral ischemia in rats. A middle cerebral artery occlusion (MCAO)/reperfusion rat model was used. The rats were sacrificed 22 h after reperfusion, and the infarct volume was evaluated. In study A, HBO (2.0 ATA), NBO (100% oxygen) and normobaric air (NBA) were each administered for 60 min in five different rat groups. The sizes of the infarcts after HBO and NBO applied during ischemia were 8.8 +/- 2.8% and 22.8 +/- 3.7% respectively of the ipsilateral non-occluded hemisphere. The infarct size after HBO applied during ischemia was statistically smaller than for NBO and NBA exposure (p < 0.01). In study B, cerebral pO(2) was measured before and after MCAO and HBO exposure (2.0 ATA for 60 min) in six rats using electron paramagnetic resonance (EPR) oximetry. The pO(2) in the ischemic hemisphere fell markedly following ischemia, while the pO(2) in the contralateral hemisphere remained within the normal range. Measurements of the pO(2) performed minutes after HBO exposure did not show an increase in the ischemic or normal hemispheres. The mean relative infarct size was consistent with the changes observed in study A. These data confirm the neuroprotective effects of HBO in cerebral ischemia and indicate that in vivo EPR oximetry can be an effective method to monitor the cerebral oxygenation after oxygen therapy for ischemic stroke. The ability to measure the pO(2) in several sites provides important information that should help to optimize the design of hyperoxic therapies for stroke.

High spatial resolution multisite EPR oximetry of transient focal cerebral ischemia in the rat.

In vivo electron paramagnetic resonance (EPR) spectroscopy can provide direct noninvasive, continuous, and repeatable measurements of oxygen in tissues. High-spatial-resolution multisite (HSRMS) oximetry is an EPR technique that uses applied magnetic field gradients to extend this capability to multiple implanted probes within the sample and accurately to estimate their respective local pO(2) values. These capabilities are crucial in experiments in which pO(2) varies across space and time and in which information about these variations is needed to describe physiologic and pathophysiologic phenomena and evaluate their responses to interventions such as therapy. One important application is the investigation of transient focal ischemia in the rat brain and the effects of treatment with hyperoxygenation. We used HSRMS oximetry with overmodulation to measure brain tissue oxygenation in a rat stroke model using lithium phthalocyanine as the oxygen probe. Oxygen measurements were made in a small cohort of rats at four implant sites during ischemia and reperfusion after transient focal ischemia initiated by occlusion of the middle cerebral artery. These measurements demonstrate the capabilities of the HSRMS oximetry technique and set the stage for more extensive physiologic studies.

The effects of Efaproxyn (efaproxiral) on subcutaneous RIF-1 tumor oxygenation and enhancement of radiotherapy-mediated inhibition of tumor growth in mice.

Efaproxiral, an allosteric modifier of hemoglobin, reduces hemoglobin-oxygen binding affinity, facilitating oxygen release from hemoglobin, which is likely to increase tissue pO(2). The purpose of this study was to determine the effect of efaproxiral on tumor oxygenation and growth inhibition of RIF-1 tumors that received X radiation (4 Gy) plus oxygen breathing compared to radiation plus oxygen plus efaproxiral daily for 5 days. Two lithium phthalocyanine (LiPc) deposits were implanted in RIF-1 tumors in C3H mice for tumor pO(2) measurements using EPR oximetry. Efaproxiral significantly increased tumor oxygenation by 8.4 to 43.4 mmHg within 5 days, with maximum increases at 22-31 min after treatment. Oxygen breathing alone did not affect tumor pO(2). Radiation plus oxygen plus efaproxiral produced tumor growth inhibition throughout the treatment duration, and inhibition was significantly different from radiation plus oxygen from day 3 to day 5. The results of this study provide unambiguous quantitative information on the effectiveness of efaproxiral to consistently and reproducibly increase tumor oxygenation over the course of 5 days of treatment, modeling the clinical use of efaproxiral. Also, based on the tumor growth inhibition, the study shows the efaproxiral-enhanced tumor oxygenation was radiobiologically significant. This is the first study to demonstrate the ability of efaproxiral to increase tumor oxygenation and to increase the tumor growth inhibition of radiotherapy over 5 days of treatment.

In Vivo EPR For Dosimetry.

As a result of terrorism, accident, or war, populations potentially can be exposed to doses of ionizing radiation that could cause direct clinical effects within days or weeks. There is a critical need to determine the magnitude of the exposure to individuals so that those with significant risk have appropriate procedures initiated immediately, while those without a significant probability of acute effects can be reassured and removed from the need for further consideration in the medical/emergency system. In many of the plausible scenarios there is an urgent need to make the determination very soon after the event and while the subject is still present. In vivo EPR measurements of radiation-induced changes in the enamel of teeth is a method, perhaps the only such method, which can differentiate among doses sufficiently for classifying individuals into categories for treatment with sufficient accuracy to facilitate decisions on medical treatment. In its current state, the in vivo EPR dosimeter can provide estimates of absorbed dose with an error approximately +/- 50 cGy over the range of interest for acute biological effects of radiation, assuming repeated measurements of the tooth in the mouth of the subject. The time required for acquisition, the lower limit, and the precision are expected to improve, with improvements in the resonator and the algorithm for acquiring and calculating the dose. The magnet system that is currently used, while potentially deployable, is somewhat large and heavy, requiring that it be mounted on a small truck or trailer. Several smaller magnets, including an intraoral magnet are under development, which would extend the ease of use of this technique.

Experimental Procedures for Sensitive and Reproducible In Situ EPR Tooth Dosimetry.

In vivo electron paramagnetic resonance (EPR) tooth dosimetry provides a means for non-invasive retrospective assessment of personal radiation exposure. While there is a clear need for such capabilities following radiation accidents, the most pressing need for the development of this technology is the heightened likelihood of terrorist events or nuclear conflicts. This technique will enable such measurements to be made at the site of an incident, while the subject is present, to assist emergency personnel as they perform triage for the affected population. At Dartmouth Medical School this development is currently being tested with normal volunteers with irradiated teeth placed in their mouths and with patients who have undergone radiation therapy. Here we describe progress in practical procedures to provide accurate and reproducible in vivo dose estimates.

Increased oxygenation of intracranial tumors by efaproxyn (efaproxiral), an allosteric hemoglobin modifier: In vivo EPR oximetry study.

PURPOSE: To determine quantitatively the changes in oxygenation of intracranial tumors induced by efaproxiral, an allosteric hemoglobin modifier. Efaproxiral reduces hemoglobin-oxygen binding affinity, which facilitates oxygen release from hemoglobin into surrounding tissues and potentially increases the pO(2) of the tumors. METHODS AND MATERIALS: The study was performed on 10 male Fisher 344 rats with 9L intracranial tumors. Electron paramagnetic resonance (EPR) oximetry was used to measure quantitatively the changes in the pO(2) in the tumors. Lithium phthalocyanine (LiPc) crystals were implanted in the tumors and in the normal brain tissue in the opposite hemispheres. We monitored the cerebral pO(2) starting 7 to 10 days after the tumor cells were implanted. NMR imaging determined the position and size of tumor in the brain. After an initial baseline EPR measurement, efaproxiral (150 mg/kg) was injected intravenously over 15 minutes, and measurements of tumor and normal brain oxygen tension were made alternately at 10-minute intervals for the next 60 minutes; the procedure was repeated for 6 consecutive days. RESULTS: Efaproxiral significantly increased the pO(2) of both the intracranial tumors and the normal brain tissue on all days. The maximum increase was reached at 52.9 to 59.7 minutes and 54.1 to 63.2 minutes after injection, respectively. The pO(2) returned to baseline values at 106 to 126.5 minutes after treatment. The maximum tumor and normal tissue pO(2) values achieved after efaproxiral treatment from Day 1 through Day 6 ranged from 139.7 to 197.7 mm Hg and 103.0 to 135.9 mm Hg, respectively. The maximum increase in tumor tissue pO(2) values from Day 2 to Day 5 was greater than the maximum increase in normal tissue pO(2). CONCLUSION: We obtained quantitative data on the timing and extent of efaproxiral-induced changes in the pO(2) of intracerebral 9L tumors. These results illustrate a unique and useful capability of in vivo EPR oximetry to obtain repeated noninvasive measurements of tumor oxygenation over a number of days. The information on the dynamics of tumor pO(2) after efaproxiral administration illustrates the ability of efaproxiral to increase intracranial tumor oxygenation.

Cerebral tissue oxygenation in reversible focal ischemia in rats: multi-site EPR oximetry measurements.

Multi-site electron paramagnetic resonance (EPR) oximetry was used in vivo to measure the partial pressure of oxygen (pO2) in reversible focal ischemia in rats. The cerebral tissue pO2 was measured simultaneously and continuously at two sites on the ischemic side and one on the normal side of the brain in the same animal prior to and at several time points after ischemia and reperfusion. The O2 at the three different sites in brain was stable over 30 min of baseline measurements. During the first 120 min of ischemia, statistically significant decreases in brain pO2 from baseline were consistently observed in the ischemic core and perifocal area. The mean values varied during the 120 min of ischemia. Reperfusion resulted in an immediate increase in PO2, but there were no significant differences between the sites over time. The result of this study seems promising for the study of ischemia and reperfusion. It appears that the technique can provide information on the PO2 under the experimental conditions needed for such a study. The levels of PO2 that occurred in these experiments are readily resolvable by multi-site EPR oximetry. In addition, the ability simultaneously to measure the pO2 in several sites provides important additional information that should help to differentiate between changes in the PO2 due toglobal or local mechanisms.

"Distant spin trapping": a method for expanding the availability of spin trapping measurements.

The technique of spin trapping is used to study a wide range of free radicals in various systems, including those generated in vitro and in vivo. But unfortunately, EPR spectrometers are not always immediately accessible at the site of experimentation, and therefore it is important to find a method that can preserve a radical adduct over longer periods of time. We describe here an alternative method in which the samples can be frozen and transported for EPR measurements at another site. Various spin adducts of DEPMPO were frozen and measured at 0 degrees C at various intervals after freezing to determine their stability in the frozen state. The radical adducts were generated by established methods and stored at two different temperatures; -196 degrees C (liquid nitrogen) and -80 degrees C (dry ice). The experiments were carried out in an aqueous solution with and without a model of reducing environment (2 mM ascorbate). The results indicate that it is feasible to store and transport spin adducts for subsequent analysis. We conclude that this approach, which we term "distant spin trapping", makes it feasible to transport samples to another site for EPR measurements. This should significantly expand the ability to use spin trapping in biology and medicine.

Modeling of the response of ptO2 in rat brain to changes in physiological parameters.

It is known that oxygen tension in tissue (ptO2) will change in response to an alteration of physiological parameters including: pCO2 in arterial blood, blood flow, capillary density, oxygen carrying capacity, and p50 of hemoglobin. We have used modeling to compute the change of PtO2 in response to changes of each physiological parameter and related these changes to experimental data. The oxygen distribution in a Krogh cylinder was computed assuming a linear decrease of hemoglobin saturation from the arterial to the venous end of the capillary. Parameters of the model were used to compute the baseline cerebral PtO2 expressed as the mean value of the PtO2 over the whole cylinder. These parameters were adjusted to derive PtO2 values close to those measured at the relevant experimental conditions. Then each desired parameter was varied to calculate the change in PtO2 related to this parameter. Effects of different factors on cerebral PtO2 were modeled and compared with experimental values obtained with various experimental interventions including: changing CBF, modifying p50 with the allosteric modifier RSR13, modification of capillary density, and hemoglobin content. An acceptable agreement of the computed and the experimental changes of the cerebral PtO2 was obtained for these experimental conditions.

Black magic and EPR oximetry: from lab to initial clinical trials.

EPR oximetry is a technique that can make repeated non-invasive measurements of the PO2 in tissues. To extend the application of EPR oximetry to humans, India ink is the probe of choice because appropriate India inks have EPR signals whose line widths are sensitive to changes in oxygen concentrations, and, most importantly, India ink already has been used extensively in humans as a marker in the skin, lymphatics, various organs during surgery, tumors, and for decoration as tattoos. We have developed an India ink that has good sensitivity to oxygen, high stability in tissues, good signal intensity, and minimal toxicity. In this article we describe the various properties of this India ink, results obtained from our animal experiments, and our first preliminary clinical results, which are part of the first systematic clinical use of EPR oximetry. The clinical results indicate that it is possible to do repeated measurements over several months and probably years after the injection of the ink, indicating that long-term follow-up studies are feasible. We are very encouraged with these results and are confident that EPR oximetry using India ink will be a non-invasive, fast, and reliable technique for pO2 measurements in clinical studies.