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The Majorana Collaboration is building the Majorana Demonstrator, a 60 kg array of high purity germanium detectors housed in an ultra-low background shield at the Sanford Underground Laboratory in Lead, SD. The Majorana Demonstrator will search for neutrinoless double-beta decay of 76Ge while demonstrating the feasibility of a tonne-scale experiment. It may also carry out a dark matter search in the 110 GeV/c2 mass range. We have found that customized Broad Energy Germanium (BEGe) detectors produced by Canberra have several desirable features for a neutrinoless double-beta decay experiment, including low electronic noise, excellent pulse shape analysis capabilities, and simple fabrication. We have deployed a customized BEGe, the Majorana Low-Background BEGe at Kimballton (MALBEK), in a low-background cryostat and shield at the Kimballton Underground Research Facility in Virginia. This paper will focus on the detector characteristics and measurements that can be performed with such a radiation detector in a low-background environment. © 2010 Elsevier B.V.

The combination of intravital microscopy and animal models of disease has propelled studies of disease mechanisms and treatments. However, many disorders afflict tissues inaccessible to light microscopy in live subjects. Here we introduce cellular-level time-lapse imaging deep within the live mammalian brain by one- and two-photon fluorescence microendoscopy over multiple weeks. Bilateral imaging sites allowed longitudinal comparisons within individual subjects, including of normal and diseased tissues. Using this approach, we tracked CA1 hippocampal pyramidal neuron dendrites in adult mice, revealing these dendrites' extreme stability and rare examples of their structural alterations. To illustrate disease studies, we tracked deep lying gliomas by observing tumor growth, visualizing three-dimensional vasculature structure and determining microcirculatory speeds. Average erythrocyte speeds in gliomas declined markedly as the disease advanced, notwithstanding significant increases in capillary diameters. Time-lapse microendoscopy will be applicable to studies of numerous disorders, including neurovascular, neurological, cancerous and trauma-induced conditions.

The authors present two cases of agenesis of the internal carotid artery (ICA) discovered incidentally on magnetic resonance imaging and confirmed on computed tomography, magnetic resonance angiography, and conventional angiography. They also propose a clinical algorithm for the workup of patients with suspected absence of the ICA.

Purpose: The goal of this work was to describe the clinical and imaging features of thoracopancreatic fistula, a rare complication of pancreatitis., Method: Nine cases of thoracopancreatic fistula proved by thoracentesis, endoscopic retrograde cholangiopancreatography (ERCP), or surgery were retrospectively and independently reviewed by two abdominal radiologists. All available imaging examinations [chest radiographs = 9, CT = 9, MR and MR cholangiopancreatography (MRCP) = 2, and ERCP = 6] were analyzed, and findings were recorded on a standardized datasheet. Available medical records (n = 7) were reviewed to determine the clinical presentation of the patients and thoracentesis results., Results: Seven of the nine patients presented with pulmonary symptoms such as dyspnea or cough. Of the seven patients with pleural fluid analysis, all demonstrated elevated amylase levels (mean 13,007 U/L). Imaging examinations revealed pancreaticopleural fistulas in six patients, a mediastinal pseudocyst in one patient, and both a pancreaticopleural fistula and a mediastinal pseudocyst in two patients. Chest radiography showed pleural fluid collections in eight patients. CT demonstrated a fluid-containing fistula in all nine patients. MR and MRCP depicted a fistula extending from the abdomen to the pleural space in the two patients with MR correlation. ERCP showed pancreatic ductal changes characteristic of chronic pancreatitis in the six patients with ERCP correlation but failed to demonstrate the fistula in two of the six patients., Conclusion: The CT, MR, MRCP, or ERCP finding of a fluid-filled tract extending from the pancreas to the thorax is characteristic of a thoracopancreatic fistula, particularly when identified in a patient who presents with pulmonary symptoms and a history of chronic pancreatitis.

Magnetic resonance (MR) cholangiography is a fast, accurate, noninvasive alternative to endoscopic retrograde cholangiography (ERC) in the evaluation of biliary tract disease. Technical improvements in imaging sequences (eg, half-Fourier rapid acquisition with relaxation enhancement) and use of phased-array coils allow high-quality imaging comparable to that available with ERC. In choledocholithiasis, common bile duct stones as small as 2 mm can be detected with MR cholangiography and appear as low-signal-intensity foci within the high-signal-intensity bile. MR cholangiography may help establish the diagnosis of malignant obstruction and is useful in the evaluation of patients in whom ERC was unsuccessful or incomplete. The role of MR cholangiography in the evaluation of intrahepatic duct disease continues to evolve. MR cholangiography plays a crucial role in evaluating postsurgical biliary tract alterations and can be used to demonstrate a variety of congenital anomalies of the biliary tract (eg, aberrant ducts, choledochal cysts, pancreas divisum). In addition, intentional or incidental imaging of the gallbladder with MR cholangiography can be used to identify calculi or help determine the presence and extent of neoplastic disease.

Because intraoperative sonography displays segmental anatomy, allows discovery of more lesions than previously suspected from preoperative imaging, surgical inspection, or palpation, and permits more certain diagnosis of problematic masses, it facilitates surgical decision-making when liver resection or cryoablation is anticipated. Intraoperative sonography provides a guidance modality to accurately place cryosurgery probes in liver masses. More precise treatment of metastatic and primary tumors of the liver is possible with cryoablation because intraoperative sonography provides a means of monitoring the growth of the enlarging freeze zone to insure adequate surgical margins. Postoperative detection of acute complications after cryosurgery is best done with computed tomography. Normally cryosurgery defects shrink with time and lose the peripheral contrast opacification seen after surgery. Gas collections, seen as a result of tissue necrosis, must be discriminated from infection. Tumor recurrence can be detected well with computed tomography or magnetic resonance imaging following hepatic cryosurgery.

A study was undertaken to determine if humans excreted pentobarbital N-glucosides as urinary metabolites following oral administration of pentobarbital. (1'RS,5RS)-1-(beta-D-Glucopyranosyl)pentobarbital ((1'RS,5RS)-PTBG) was isolated from the urine of one subject. The two diastereomers, (1'RS,5R)-PTBG and (1'RS,5S)-PTBG were separated and found to be identical to synthetic standards when compared using HPLC retention times coupled with UV (with and without post-column ionization) and mass spectrometry (HPLC/MS). A HPLC method was developed for detecting and quantifying (1'RS,5R)-PTBG, (1'RS,5S)-PTBG and pentobarbital in urine. Following a single oral dose of sodium pentobarbital to male subjects (n = 6), 1.6-6.2% of the pentobarbital dose was excreted as (1'RS,5S)-PTBG over 60 hours. (1'RS,5R)-PTBG was also detected in one subject and accounted for 0.3% of the pentobarbital dose. Using a modified HPLC system, the four pentobarbital N-glucosides were resolved and analysis of a partially purified pentobarbital N-glucoside extract from one subject indicated that only (1'R,5R)-PTBG and (1'S,5S)-PTBG could be detected as urinary excretion products. These results indicate that the side chain chirality of pentobarbital may influence the observed enantioselectivity for the formation and/or urinary excretion of the pentobarbital N-glucosides.

In vitro translation studies indicate that the beta 2-microglobulin (beta 2-m) light chain influences the formation of intrachain disulfide bonds in class I major histocompatibility complex (MHC) molecules during their biosynthesis. We now have examined the influence of beta 2-m on class I MHC intrachain disulfide bond formation in vivo. Using beta 2-m+ and beta 2-m- derivatives of a cell line transfected with the mouse H-2Ld gene, we show that all of the H-2Ld molecules from beta 2-m+ cells have both the alpha 2 and alpha 3 intrachain disulfide bonds, whereas about 50% of the H-2Ld molecules from beta 2-m- cells have only one of these bonds. All of the free H-2Ld heavy chains from beta 2-m+ cells can undergo a peptide-induced conformational change and can bind exogenous peptide and beta 2-m stably in vitro. Only those H-2Ld molecules from beta 2-m- cells, which have both intrachain disulfide bonds, undergo a peptide- and beta 2-m-induced conformational change in vitro. These H-2Ld molecules do not bind beta 2-m and peptide stably in vitro. From these results emerges a greater understanding of the role of beta 2-m at the time of class I MHC molecule synthesis: beta 2-m promotes intrachain disulfide bond formation in the class I MHC molecule and additionally affects class I MHC structure to render it competent to form stable trimolecular complexes with peptide and beta 2-m.

Recent studies have shown that the endoplasmic reticulum (ER)-resident protein, calnexin, associates with class I major histocompatibility complex (MHC) molecules early in their biosynthesis. It has been suggested that calnexin participates in the assembly of class I MHC molecules or in the retention within the ER of unassembled class I molecules. We have examined the role of phosphorylation of calnexin in its association with mouse class I MHC molecules. We show that phosphocalnexin associates with H-2Ld and H-2Db molecules but not with H-2Kb and H-2Dd molecules, although calnexin-H-2Kb association can be demonstrated. These observations are interesting in light of the fact that H-2Kb and H-2Dd molecules are transported out of the ER more rapidly than are H-2Ld and H-2Db molecules. H-2Ld and H-2Db molecules differ in amino acid sequence only in their membrane-distal alpha 1 and alpha 2 domains. Nevertheless, the affinity of phosphocalnexin for H-2Ld is greater than its affinity for H-2Db. Furthermore, H-2Db becomes endoglycosidase H-resistant more slowly in cells in which it associates with phosphocalnexin than in cells in which it does not. Ca2+ ionophore A23187 prevents association of phosphocalnexin with H-2Ld molecules in vivo but does not cause the disruption of phosphocalnexin-H-2Ld complexes after they have formed. A23187 does not prevent assembly of H-2Ld-beta 2-microglobulin (beta 2-m) heterodimers. Furthermore, phosphocalnexin is found associated with H-2Ld molecules regardless of their state of assembly with beta 2-m and antigenic peptide. These results suggest that phosphocalnexin association with class I MHC molecules does not play a role in assembly of the class I MHC-beta 2-m-peptide complex nor in preventing release of unassembled class I molecules from the ER but may otherwise influence their rate of transport through the ER.

An antiserum was generated against a synthetic peptide corresponding to a portion of the cytoplasmic domains of the H-2Ld and H-2Db class I major histocompatibility complex molecules of the mouse. This antibody preparation, R4, binds exclusively to endoglycosidase H-resistant H-2Ld/Db molecules which are not associated with beta 2-microglobulin. Interestingly, acquisition of resistance to endoglycosidase H precedes acquisition of R4 reactivity by 30 min. R4-reactive H-2Ld and H-2Db molecules occur on the cell surface and are phosphorylated in vivo. Other studies show that the tyrosine in the cytoplasmic domain is accessible to radioiodination on only a subset of H-2Ld molecules, and that the two-dimensional electrophoretic profiles of phosphorylated H-2L/Db molecules, of R4-reactive molecules, and of H-2Ld molecules radiolabeled on this cytoplasmic domain tyrosine are virtually identical. R4-reactive H-2Ld molecules do not undergo the peptide- and beta 2-microglobulin-induced conformational changes characteristic of free class I major histocompatibility complex heavy chains. The accessibility of the H-2Ld cytoplasmic domain to R4 and to radioiodination late in biosynthesis and its biological significance are discussed.

L-Lactate dehydrogenase (LD) catalyzes the interconversion of pyruvate and lactate. Using a spectrophotometric assay to determine LD activity, incubation of rabbit, porcine, and bovine LD-1 and LD-5 isozymes with the protease subtilisin (Carlsberg) gave first-order degradation kinetics. Degradation half-lives were significantly lower for the LD-5 isozymes from the three species when incubated with subtilisin at temperatures from 4 degrees C to 25 degrees C. The energy involved in the degradation process, however, was not different. The activation energy for the conversion of pyruvate to lactate by LD-1 at pH 7.4 was significantly higher than that for LD-5 for all three species examined (P less than 0.005). Thermocalorimetry showed that the LD-1 isozymes have both a higher mean temperature of denaturation and a higher heat uptake during the denaturation process than corresponding LD-5 forms. The results suggest that the LD-5 isozymes in the species studied are more metabolically efficient, whereas the LD-1 forms have greater structural stability.

Protein phosphorylation is widely believed to play a regulatory role in signal transduction, mitosis, cell proliferation, cell motility, cell shape, gene regulation, and many other cellular processes. Thus, the quantitation of phosphorylation of specific cellular proteins may provide insight into the mechanisms by which phosphorylation is employed in regulation. Moreover, identification of phosphorylation substrates of various cellular kinases provides an important first step in determining their role in cellular regulation. However, accurate measurement of the differential phosphorylation of cellular proteins under different physiological conditions is often difficult to achieve. To address this problem, we have developed an in vivo double-labeling protocol (utilizing [3H]-, [14C]-, or [35S]-radiolabeled amino acids and [32P]-orthophosphate) that allows the quantitation of the amount of specific phosphorylation of a given protein from densitometric analysis of autoradiograms of polyacrylamide gels. This double-labeling strategy provides a means of quantitating the phosphorylation of individual biosynthetically labeled proteins. This method can be used in the analysis of immunoprecipitated proteins, proteins from subcellular fractions, such as nuclei or selected membrane fractions, or even total cellular proteins displayed on two-dimensional gels.