SU-E-I-101: Initial Implementation and Evaluation of AAPM TG-150 Draft Image Receptor Non-Uniformity Testing Recommendations.

PURPOSE: To implement in software the procedures described in AAPM Task Group 150's draft recommendations for image receptor performance testing, and to evaluate the effectiveness and practicality of these procedures. METHODS: Images of flat fields were acquired using digital x-ray image receptors at 6 cooperating institutions. Four flat field images obtained with each detector spanned a range of input detector air kerma. Software based on AAPM TG150's draft report processed the test images and generated results. Image receptor response and several measures of non-uniformity were evaluated. Images were divided into 10 mm square regions, after eliminating 10 mm borders. For each region, signal (mean), noise (standard deviation) and SNR were calculated. Characteristic signal, noise and SNR were calculated based on average values from all regions. Local non-uniformity for signal (SLN), noise (NLN) and SNR (SNRLN) were expressed as the maximum ratio of the absolute difference between each region's value and its 4 nearest neighbors, to the respective characteristic value. Global non-uniformity (SGN, NGN, SNRGN) were expressed similarly but differences between maximum and minimum values obtained from the regions were used (without comparison to local neighbors). RESULTS: TG150 tests discriminated between good and poorly performing detectors. Improper detector calibration was detectable, with noise non-uniformity proving to be a more sensitive measure than signal or SNR non-uniformity. Detector rotation relative to calibration conditions produced a greater change in signal non-uniformity than the other measures. Image receptor structured noise was characterized by an increase in noise non-uniformity with incident air kerma. CONCLUSIONS: AAPM TG150's proposed approach to image receptor testing was implemented and evaluated. The approach appears to be an effective and practical one for routine quality assurance testing of digital radiographic image receptors.

SU-E-I-103: Detecting Anomalous Pixels and Correlated Artifacts in Digital Detectors from Flat-Field Images.

PURPOSE: Anomalous pixels may be defined as those pixels whose exposure response relationship is deviant from the typical, expected or calibrated response. A group of anomalous pixels may Result in visible correlated artifacts. Here we demonstrate an approach to identify anomalous pixels and correlated artifacts using flat-field images. METHODS: Using manufacturer specific calibration geometry, sets of four flat-field images per detector were obtained with varying input air kerma values (0.5 to 160 µGy) from 9 digital detectors at 6 institutions. Images obtained before and after calibration, with both proper and improper gain maps and structured artifacts were additionally acquired with some detectors. Image analysis methodology under consideration by AAPM Task Group 150 was used.After eliminating 10mm borders, images were divided into square regions (100mm2 ). Anomalous pixels were identified as pixels within each region with valuesabove or below ±3 standard deviations (SD) relative to the mean value of the region. If these pixels were identified in all four images comprising a set, then they were reported as anomalous. Line artifacts were identified as rows and columns with cumulative profile values that were above or below ±3 SD with respect to the mean value of neighboring profiles in the set of four flat-field mages. Results were verified with visual inspection of the images. RESULTS: For four sets of images, the algorithm did not identify any anomalous pixels, and none were spotted on visible inspection as well, while for five sets of images the identified anomalous pixels matched visual inspection results. Anomalous pixel detection failed in regions with an unusually large number of defects and structured noise, since those regions exhibited relatively large SD. Line artifacts consistent with visual analysis were identified correctly when present. CONCLUSIONS: A practical approach to identify anomalous pixels and correlated artifacts from flat-field images is demonstrated.

Preferential amplification is minimised in long-PCR systems.

The advent of long PCR (XL-PCR) has proven to be a major advance in PCR technology and is currently being utilised to investigate numerous biological systems. The analysis of mixed DNA populations is a particularly useful application for XL-PCR. For example, XL-PCR has been used to investigate the occurrence of heterogeneous mitochondrial DNA (mtDNA) rearrangement mutations. With XL-PCR it became possible to amplify the entire length of the mtDNA chromosome and detect any mtDNA deletion or insertion mutations based on a measurable change in overall sequence length. In the present communication, XL-PCR and conventional short-length PCR were used to amplify mitochondrial DNA sequences from several human vastus lateralis skeletal muscle samples. The experiments demonstrated that there was minimal preferential amplification of shorter DNA sequences with XL-PCR and was significantly less than the preferential amplification of shorter sequences observed with conventional PCR. Also, XL-PCR amplification of the complete mtDNA sequence from control DNA containing a single mtDNA template (leucocyte extracts) showed that the generation of PCR artefacts was not a predisposed failing of the system but was dependant on the standard rules that govern the set up and optimisation of any PCR reaction. In optimised systems, XL-PCR artefacts were not generated and a single PCR product was always recovered.

Gastrointestinal imaging: a systems analysis comparing digital and conventional techniques.

OBJECTIVE: The purpose of this study was to compare digital and conventional methods of gastrointestinal imaging based on the cost of image storage and estimated overall costs, radiation exposure to the patient, and duration of the examination. MATERIALS AND METHODS: Our study sample consisted of 128 patients who underwent conventional gastrointestinal studies (64 double-contrast upper gastrointestinal examinations and 64 double-contrast barium enemas) and 139 patients who underwent digital gastrointestinal studies (66 double-contrast upper gastrointestinal examinations and 73 double-contrast barium enemas). The number of images and films for each study was recorded, and the mean cost of image storage and the estimated overall costs for digital versus conventional studies were calculated. Both the duration of fluoroscopy and the time from start to completion of the study were obtained from our radiology information system. From these data, we calculated mean radiation exposure to the patient and the duration of the examination. Finally, referring physicians completed a questionnaire about their level of satisfaction with paper prints generated from digital gastrointestinal studies. RESULTS: When digital studies were compared with conventional studies, the mean cost of image storage decreased by 45% and the estimated overall 10-year costs decreased by 8%. The mean number of spot images increased by 8% for upper gastrointestinal examinations and by 25% for barium enema examinations, whereas the mean duration of fluoroscopy decreased by 4% and by 10%, respectively. As a result, radiation exposure to patients increased by only 2%, a difference that did not approach statistical significance. Finally, the mean duration of examinations decreased by 24% for upper gastrointestinal examinations and by 33% for barium enemas. Approximately 85% of the physicians who completed the questionnaires indicated that they reviewed the paper prints generated from digital studies and that they would like to continue receiving them. CONCLUSION: Digital gastrointestinal imaging systems are associated with higher initial costs than conventional systems, but the long-term costs of these digital imaging systems are slightly less because of the lower cost of image storage, and radiation exposure to patients is comparable. The shorter duration of digital examinations is a potential benefit of this technology, allowing improved patient throughput. Finally, referring physicians have a high level of satisfaction with paper prints generated from digital imaging.

Modulation of Fos-mediated AP-1 transcription by the promyelocytic leukemia protein.

The growth and transformation suppressor function of promyelocytic leukemia (PML) protein are disrupted in acute promyelocytic leukemia (APL) as a result of its fusion to the RARalpha gene by t(15;17) translocation. There is significant sequence homology between the dimerization domain of PML and the Fos family of proteins, which imply that PML may be involved in AP-1 activity. Here we show that PML can cooperate with Fos to stimulate its AP-1-mediated transcriptional activity. Cotransfection of PML with GAL4/Fos strongly induced Fos-mediated activation of GAL4-responsive reporters, indicating a functional interaction between Fos and PML in vivo. Deletion analysis of Fos and PML demonstrated that the intact C-terminal domain of Fos (containing the dimerization domain), and the RING-finger, B1 box and nuclear localization domains of PML are involved in the cooperative activity of Fos and PML. Immunoprecipitation and electrophoretic mobility shift assay showed that PML is associated with the AP-1 complex. PMLRARalpha was also found to enhance the transcriptional activity of GAL4/Fos. The addition of retinoic acid abrogated the PMLRARalpha, but not PML-induced stimulation of GAL4/Fos activity in a dose-dependent manner. This study demonstrated that PML is involved in the AP-1 complex and can modulate Fos-mediated transcriptional activity, which may contribute to its growth suppressor function.

The age-associated decrease in the amount of amplifiable full-length mitochondrial DNA in human skeletal muscle.

There has been a continuous evolution in our concept [1] that mtDNA undergoes a range of mutations with age and that such alterations lead to a decline in mitochondrial bioenergy capacity. Here we report that a wide range of deletion mutations accumulate with age and the amount of full-length mtDNA (FLmtDNA) amplifiable by extra-long PCR (XL-PCR) markedly decreases with age. An analysis of single human quadriceps muscle fibres reveals a close correlation between the decrease in FLmtDNA and the decline in cytochrome c oxidase activity, an exemplifier of mitochondrial bioenergy. However, Southern blotting analysis of unamplified genomic DNA shows that there is little decrease in FLmtDNA in aged quadriceps. The results are interpreted to indicate that while there is little change in the total mtDNA with age, nonetheless a significant proportion of this mtDNA is extensively damaged such that it cannot be amplified by XL-PCR. The amplifiable FLmtDNA, which putatively represents the functional component of the mtDNA, decreases markedly with age.

The universality of bioenergetic disease. Age-associated cellular bioenergetic degradation and amelioration therapy.

During the present century there has been a dramatic change in life expectancy in advanced societies, now exceeding 80 years. As distinct from life expectancy, life potential is said to be at least 120 years, so that the continuing increase in knowledge has the potential for further major changes in the survival of humans conceivably in the near future. This presentation will be concerned with one aspect of the development of biomedical advances related in part to a concept of an "age-related universality of bioenergetic disease," and its potential amelioration and proposed impact on age-related disease and lifestyle. Aging is a complex biological process associated with a progressive decline in the physiological and biochemical performance of individual tissues and organs, leading to age-associated disease and senescence. Consideration of the progressive accumulation of mitochondrial DNA mutation with age and the tissue/cellular bioenergy decline associated with the aging process has led us to the proposal of a "universality of bioenergetic disease" and the potential for a redox therapy for the condition. This concept envisages that a tissue-bioenergetic decline will be intrinsic to various diseases of the aged and thereby contribute to their pathology, in particular, heart failure, degenerative brain disease, muscle and vascular diseases, as well as other syndromes. The information and concepts embodied in this proposal will be reviewed under the following headings: (1) mitochondrial DNA deletion mutation in some tissue is very extensive and shows mosaicism; (2) age-associated tissue/cellular bioenergy mosaic closely corresponds to the mtDNA profile; (3) cellular bioenergy as a function of mitochondrial bioenergy, glycolysis, and plasma membrane oxidoreductase; (4) redox therapy for the reenergization of cells, tissues, and whole organs. A redox therapy based on coenzyme Q10 has demonstrated profound alteration in heart function of old rats; no significant effect was observed with young rats.

Transcriptional repression by the promyelocytic leukemia protein, PML.

Acute promyelocytic leukemia is characterized by the presence of a t(15; 17) chromosomal translocation which results in the expression of a chimeric gene product, PMLRAR alpha, consisting of an N-terminal-truncated retinoic acid receptor-alpha fused to a C-terminal-truncated PML. Several structural features, and regions of homology to known transcription factors, suggest that PML may be involved in the regulation of gene expression. In this study we have analyzed the transcriptional regulatory activity of PML using chimeric GAL4/PML constructs and GAL4-responsive reporter plasmids. The data presented demonstrate that PML, when fused to the DNA-binding domain of GAL4 (GAL4/PML), inhibits transcription from GAL4-responsive promoters. The magnitude of this repression is cell type and promoter dependent, and deletion studies show that the putative coiled-coil and part of the serine-rich regions of PML are required for this activity. These regions are also shown to be responsible for the repression of transcription activity from the EGFR promoter. The data presented also demonstrate that GAL4/PML can recruit PMLRAR alpha resulting in the retinoid-inducible transcriptional activation of a GAL4-responsive promoter, a function dependent on the presence of the coiled-coil region of PMLRAR alpha.

Contrast and dose with Mo-Mo, Mo-Rh, and Rh-Rh target-filter combinations in mammography.

PURPOSE: To study the effect of three mammography target-filter combinations on contrast and dose. MATERIALS AND METHODS: With screen-film sensitometry, the contrast of a calcification target embedded in simulated breast tissue was measured for three target-filter combinations--molybdenum-molybdenum (Mo-Mo), molybdenum rhodium (Mo-Rh), and rhodium-rhodium (Rh-Rh)--as a function of x-ray tube potential, breast thickness, and breast composition. The corresponding average glandular tissue doses were also determined. RESULTS: Contrast and dose decreased with increasing kilovolt peak with all three target-filter combinations. Contrast was highest for Mo-Mo and lowest for Rh-Rh for images exposed with a low kilovoltage (< 29 kVp). For thick or radiographically dense phantoms, the contrast produced with Mo-Mo was less than or equal to that produced by the other two x-ray spectra when a higher kilovoltage (> or = 29 kVp) was selected. Average glandular dose was greatest for Mo-Mo and lowest for Rh-Rh for all phantom thicknesses, breast compositions, and tube potentials studied. CONCLUSIONS: For the thick or dense breast, the alternative target-filter selections can achieve contrast comparable to or better than that obtainable with Mo-Mo while using a smaller dose.

Normalized average glandular dose in molybdenum target-rhodium filter and rhodium target-rhodium filter mammography.

PURPOSE: To determine the normalized glandular dose (DgN) for molybdenum target-rhodium filter (Mo-Rh) and rhodium target-rhodium filter (Rh-Rh) mammography and compare the average glandular doses (Dg) that resulted with a conventional molybdenum target-molybdenum filter (Mo-Mo) source assembly. MATERIALS AND METHODS: X-ray spectra models for Mo-Rh and Rh-Rh were developed and used to calculate DgN values for these target-filter combinations as a function of x-ray tube potential, half-value layer, and breast thickness for three breast compositions. For the average glandular dose comparisons, 50/50 phantoms were imaged for the three target-filter source assemblies at three tube potentials. RESULTS: For the same parameters, DgN values for Mo-Rh and Rh-Rh were higher than for Mo-Mo. At the same voltage, the exposures required to image breast phantoms are substantially lower, and as a result, Dgs are also less with Mo-Rh and Rh-Rh than with Mo-Mo. CONCLUSION: DgN values presented permit practical evaluations of average glandular doses for Mo-Rh and Rh-Rh mammography. At a given potential, dose savings are realized with Mo-Rh and Rh-Rh source assemblies.

Computed radiography: user-programmable features and capabilities.

Digital or computed radiography (CR) using photostimulable storage phosphor plate technology is becoming increasingly popular in certain clinical applications, such as bedside radiography, where it possesses clear advantages over conventional screen-film imaging. The majority of CR systems in clinical use have been manufactured by Fuji Medical Systems USA, Inc (Stamford, CT) and provide a surprising degree of flexibility. Fuji CR units are delivered with preset menus, hardcopy format, and image-processing parameters for each examination. Of practical importance is that users may change the exam menu and printed film format as well as the image-processing parameters for each examination. There is, however, a lack of documentation describing these features and how they are programmed. This paper addresses these issues. Examples are given on how to change: 1) the printed film format, 2) the contrast and gray-scale processing, 3) spatial frequency enhancement, and 4) the appearance of the operator interface menus.

Description of a prototype emission-transmission computed tomography imaging system.

We have developed a prototype imaging system that can perform simultaneous x-ray transmission CT and SPECT phantom studies. This system employs a 23-element high-purity-germanium detector array. The detector array is coupled to a collimator with septa angled toward the focal spot of an x-ray tube. During image acquisition, the x-ray fan beam and the detector array move synchronously along an arc pivoted at the x-ray source. Multiple projections are obtained by rotating the object, which is mounted at the center of rotation of the system. The detector array and electronics can count up to 10(6) cps/element with sufficient energy-resolution to discriminate between x-rays at 100-120 kVp and gamma rays from 99mTc. We have used this device to acquire x-ray CT and SPECT images of a three-dimensional Hoffman brain phantom. The emission and transmission images may be superimposed in order to localize the emission image on the transmission map.

Systematic bias in basis material decomposition applied to quantitative dual-energy x-ray imaging.

Basis material decomposition represents dual-energy x-ray attenuation measurements in terms of the attenuation coefficients or thickness of two standard materials which, when combined, produce attenuation equivalent to the object being measured. In tomographic imaging, the reconstructed attenuation coefficient is calculated in terms of the attenuation coefficients of the basis materials, while in projection imaging, the thicknesses of two materials can be specified in terms of the basis materials. This analysis shows that basis material decomposition is exact in a dual-monoenergetic system, but for broad spectra, x-ray beam hardening introduces a bias into quantitative measurements. The error is small enough that it can be ignored when dual-energy imaging is used primarily to enhance the contrast of one material over another. The magnitude of the error in quantitative measurements depends on the details of the specific application including the energy of the x-ray beam, and the composition and thickness of the materials included in the object. The magnitude of the error for dual-energy bone densitometry has been analyzed using a first-order propagation of error analysis and the calculations verified by computer simulation. This analysis shows that the magnitude of the systematic error can be as high as 3% for 1 g/cm2 of bone mineral when aluminum and acrylic basis materials are used for the calibration. This systematic error is eliminated when the basis materials are the same as the materials that are being quantified (i.e., bone mineral and water).

A prototype high-purity germanium detector system with fast photon-counting circuitry for medical imaging.

A data-acquisition system designed for x-ray medical imaging utilizes a segmented high-purity germanium (HPGe) detector array with 2-mm wide and 6-mm thick elements. The detectors are contained within a liquid-nitrogen cryostat designed to minimize heat losses. The 50-ns pulse-shaping time of the preamplifier electronics is selected as the shortest time constant compatible with the 50-ns charge collection time of the detector. This provides the detection system with the fastest count-rate capabilities and immunity from microphonics, with moderate energy resolution performance. A theoretical analysis of the preamplifier electronics shows that its noise performance is limited primarily by its input capacitance, and is independent of detector leakage current up to approximately 100 nA. The system experimentally demonstrates count rates exceeding 1 million counts per second per element with an energy resolution of 7 keV for the 60-keV gamma ray photon from 241Am. The results demonstrate the performance of a data acquisition system utilizing HPGe detector systems which would be suitable for dual-energy imaging as well as systems offering simultaneous x-ray transmission and radionuclide emission imaging.

The segregation of the 2 mu-based yeast plasmid pJDB248 breaks down under conditions of slow, glucose-limited growth.

The stability of the 2 mu-based yeast plasmid pJDB248 in Saccharomyces cerevisiae S150-2B(cir0) was investigated in glucose-limited chemostat culture. Plasmid-free cells were detected by loss of (plasmid-encoded) leucine prototrophy and confirmed by colony hybridization. The plasmid was considerably more stable at a high dilution rate (0.12 h-1) than at a lower dilution rate (0.05 h-1). The average plasmid copy number in the cells retaining the plasmid remained constant at approximately 50 in the high dilution rate culture whereas it rose to almost 600 in the slow dilution rate culture. However, in both cultures the overall plasmid level in the total population remained constant, indicating that plasmid segregation breaks down at the low growth rate. Similar experiments on the native 2 mu plasmid demonstrated high stability and no significant differences between the high and low growth rate cultures. It is postulated that the difference in behaviour between the native and chimeric plasmids is related to an interaction between the growth conditions and the loss of the D gene product.

A direct study of the relative synthesis of petite and grande mitochondrial DNA in zygotes from crosses involving suppressive petite mutants of Saccharomyces cerevisiae.

Work in recent years has produced indirect evidence to support the view that the phenomenon of suppressiveness in yeast is the result of the ability of the petite mtDNA to out-replicate the wild-type genome. We have developed a method, based on fluorography of gels containing restriction fragments of radioactively labelled zygotic mtDNA, by which it has been possible to follow directly the incorporation of label into the two mtDNA species and hence their relative synthesis. Four petite isolates of 70%, 43%, 23% and 12% suppressiveness were tested by this method in crosses with a grande strain. Only the mtDNA from the 70% suppressive petite showed a replicative advantage over the grande mtDNA. The mtDNA from the 43% and 23% suppressive actually appeared to undergo, if anything, less replication in the zygote than the grande mtDNA. It is concluded that while some petites may exhibit suppressiveness as a result of enhanced replicative efficiency of their mtDNA, this cannot be the explanation for all suppressive petite strains.

The use of restriction endonucleases.

The discovery of the mode of action of the class of bacterial enzymes known as restriction endonucleases provided the major breakthrough in opening up the field of genetic engineering. In vivo, these enzymes are involved in recognizing and cutting up foreign DNA entering the cell; their most likely role is thus protecting the bacteria against phage infection. The property that is relevant to us is that these enzymes recognize specific DNA sequences. The enzymes used in DNA manipulations are in fact known as Class II restriction endonucleases; these enzymes cut the DNA within the recognition sequence at a defined point. Treatment of a DNA sample with such enzymes will thus result in each molecule being cut at the same positions and thereby lead to the formation of reproducible fragments.

Bacterial transformation.

If one consults any genetics textbook written before the mid-1970s, one will find much on the transformation of such bacterial species as Bacillus subtilis, but no mention of Escherichia coli. The method now most generally used for introducing DNA into E. coli is based on a study by Mandel and Higa in 1970 (1), who demonstrated that calcium chloride treatment would greatly enhance the ability of the cells to take up naked bacteriophage λ DNA. Later it was shown that the same method would also allow uptake of other DNA species including, in particular, plasmid DNA.

Yeast transformation.

The yeast Saccharomyces cerevisiae has tremendous advantages as a host in gene cloning experiments. It is a microorganism for which most of the techniques developed in bacterial work can be applied, including chemical mutagenesis, selective plating, and replica plating. Being the basis of an ancient industry, its fermentation characteristics are well understood. It is also a eukaryote with nitotic and meiotic divisions, cellular compartmentalization, and post-translational modification of proteins similar to that seen in higher species. Yeast is thus an ideal organism to turn to in experiments for which bacterial hosts are not suitable, but microbial techniques are nonetheless required.