Calibration of mixer amplitude and phase imbalance in superconducting circuits.

An important device for modulation and frequency translation in the field of circuit quantum electrodynamics is the in-phase and quadrature mixer, an analog component for which calibration is necessary to achieve optimal performance. In this paper, we introduce techniques originally developed for wireless communication applications to calibrate upconversion and downconversion mixers. A Kalman filter together with a controllable carrier frequency offset calibrates both mixers without removing them from the embedding measurement infrastructure. These techniques can be embedded into room temperature control electronics and hopefully find widespread use as circuit quantum electrodynamics devices continue to grow in complexity.

Genetic Variability of Two Leaffooted Bugs, Leptoglossus clypealis and Leptoglossus zonatus (Hemiptera: Coreidae) in the Central Valley of California.

Leaffooted plant bugs (LFPBs) (Leptoglossus spp., Guérin-Méneville) (Hemiptera: Coreidae) are large seed-feeding bugs native to the Western Hemisphere. In California, several Leptoglossus spp. feed on almonds, pistachios, and pomegranate and are occasional pests. The objective of this study was to survey the different species of Leptoglossus present in almond, pistachio, and pomegranate orchards in the Central Valley of California. We used two molecular markers, amplified fragment length polymorphisms (AFLPs) and mitochondrial DNA COI, to determine the number of species or strains of each species, and to infer whether individuals of each species move and possibly interbreed with populations from the other host plants. Two species of leaffooted bugs were abundant, Leptoglossus clypealis Heidemann, and Leptoglossus zonatus (Dallas). L. clypealis was collected in almond and pistachio, while L. zonatus was found on all three host plants, but was the dominant species in pomegranate. The AFLP results indicated that L. clypealis consisted of one species, which suggests it moves between almonds and pistachios during the growing season. Mitochondrial DNA COI for L. clypealis found 1-2% divergence between sequences, and a high haplotype diversity of 0.979 with 17 haplotypes. The AFLP results for L. zonatus found two genetically divergent populations which were morphologically similar. The mtDNA COI sequences for L. zonatus were used for haplotype analysis; three haplotypes were found in California, with one haplotype shared with collections from Brazil. The importance of genetic variability and cryptic species for pest management are discussed.

Mouse blood monocytes: standardizing their identification and analysis using CD115.

Monocytes have been used to assess immune dysfunction and disease. While mouse models are a useful longitudinal analog, few researchers have assessed changes in mouse monocytes. The purpose of this study was to provide recommendations for the sample processing and flow cytometric analysis of mouse blood monocytes. Blood was drawn in a non-lethal manner from CD-1 male mice to be used in three experiments. Experiment 1 compared commonly used mouse monocyte markers. Experiment 2 compared the stability of CD115 expression after immediate (0h) and delayed (2 and 4h) processing following blood collection under various experimental conditions (laser strength, anticoagulant, and storage temp.). Experiment 3 compared the consistency of CD115(+) monocyte and subset concentrations using decreasing (40, 20, 10 and 5µL) volumes of blood. In experiment 1, >95% of CD115(+) events co-expressed CD11b; >85% co-expressed CD14. 70% of CD14(+) and 50% of CD11b(+) events co-expressed CD115. In experiment 2, CD115 expression decreased by 33% between 0 and 4h when stored at room temperature. Blood treated with EDTA and refrigerated maintained CD115 stability. In experiment 3, calculated concentrations for total monocyte events varied by <10% when 40, 20 and 10µL of blood were stained. While CD115 staining provides the most distinct monocyte population, it is important to treat blood with EDTA and refrigerate if sample processing will be delayed over 2h. Collectively, the findings of the present study outline important considerations that must be addressed when examining mouse monocytes in small, non-lethal blood samples.

Reconstructing nonlinearities with intermodulation spectroscopy.

We describe a method of analysis which allows for reconstructing the nonlinear disturbance of a high Q harmonic oscillator. When the oscillator is driven with two or more frequencies, the nonlinearity causes intermodulation of the drives, resulting in a complicated spectral response. Analysis of this spectrum allows one to approximate the nonlinearity. The method, which is generally applicable to measurements based on resonant detection, increases the information content of the measurement without requiring a large detection bandwidth, and optimally uses the enhanced sensitivity near resonance to extract information and minimize error due to detector noise.

Regional chemotherapy for unresectable primary liver cancer: results of a phase II clinical trial and assessment of DCE-MRI as a biomarker of survival.

BACKGROUND: This study reports the results of hepatic arterial infusion (HAI) with floxuridine (FUDR) and dexamethasone (dex) in patients with unresectable intrahepatic cholangiocarcinoma (ICC) or hepatocellular carcinoma (HCC) and investigates dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) assessment of tumor vascularity as a biomarker of outcome. PATIENTS AND METHODS: Thirty-four unresectable patients (26 ICC and eight HCC) were treated with HAI FUDR/dex. Radiologic dynamic and pharmacokinetic parameters related to tumor perfusion were analyzed and correlated with response and survival. RESULTS: Partial responses were seen in 16 patients (47.1%); time to progression and response duration were 7.4 and 11.9 months, respectively. Median follow-up and median survival were 35 and 29.5 months, respectively; 2-year survival was 67%. DCE-MRI data showed that patients with pretreatment integrated area under the concentration curve of gadolinium contrast over 180 s (AUC 180) >34.2 mM.s had a longer median survival than those with AUC 180 <34 mM.s (35.1 versus 19.1 months, P = 0.002). Decreased volume transfer exchange between the vascular space and extracellular extravascular space (-DeltaK(trans)) and the corresponding rate constant (-Deltak(ep)) on the first post-treatment scan both predicted survival. CONCLUSIONS: In patients with unresectable primary liver cancer, HAI therapy can be effective and safe. Pretreatment and early post-treatment changes in tumor perfusion characteristics may predict treatment outcome.

Phase I trial of adjuvant hepatic arterial infusion (HAI) with floxuridine (FUDR) and dexamethasone plus systemic oxaliplatin, 5-fluorouracil and leucovorin in patients with resected liver metastases from colorectal cancer.

BACKGROUND: The purpose of the study was to determine the maximum tolerated dose of systemic oxaliplatin (oxal), 5-fluorouracil (5-FU) and leucovorin (LV) that could be administered with hepatic arterial infusion (HAI) of floxuridine (FUDR) and dexamethasone (Dex) in the adjuvant setting after hepatic resection. METHODS: Thirty-five patients with resected liver metastases were entered into a phase I trial using HAI FUDR/Dex with escalating doses of oxal and 5-FU. RESULTS: The initial dose of HAI FUDR was fixed at 0.12 mg/kg x pump volume divided by pump flow rate plus Dex infused over the first 2 weeks of a 5-week cycle. Systemic chemotherapy was delivered on days 15 and 29 with the doses of oxal escalated from 85 to 100 mg/m2 and the 5-FU 48-h continuous infusion doses from 1000 to 2000 mg/m2. The LV dose was fixed at 400 mg/m). Dose-limiting toxic effects were diarrhea, 8.5%, and elevated bilirubin, 8.5%. With a median follow-up of 43 months, the 4-year survival and progression-free survival were 88% and 50%, respectively. CONCLUSIONS: Adjuvant therapy after liver resection with HAI FUDR/Dex plus systemic oxal at 85 mg/m2 and 5-FU by continuous infusion at 2000 g/m2 with LV at 400 mg/m2 is feasible and appears effective. Randomized studies comparing this regimen to systemic FOLFOX are suggested.

Spin injection and relaxation in a mesoscopic superconductor.

We study spin transport in a superconducting nanowire using a set of closely spaced magnetic tunnel contacts. We observe a giant enhancement of the spin accumulation of up to 5 orders of magnitude on transition into the superconducting state, consistent with the expected changes in the density of states. The spin relaxation length decreases by an order of magnitude from its value in the normal state. These measurements, combined with our theoretical model, allow us to distinguish the individual spin-flip mechanisms present in the transport channel. Our conclusion is that magnetic impurities rather than spin-orbit coupling dominate spin-flip scattering in the superconducting state.

Phase-charge duality of a josephson junction in a fluctuating electromagnetic environment.

We have measured the current-voltage characteristics of a single Josephson junction placed in a high impedance environment. The transfer of Cooper pairs through the junction is governed by overdamped quasicharge dynamics, leading to Coulomb blockade and Bloch oscillations. Exact duality exists to the standard overdamped phase dynamics of a Josephson junction, resulting in a dual shape of the current-voltage characteristic, with current and voltage changing roles. We demonstrate this duality with experiments which allow for a quantitative comparison with a theory that includes the effect of fluctuations due to the finite temperature of the electromagnetic environment.

Direct demonstration of decoupling of spin and charge currents in nanostructures.

The notion of decoupling of spin and charge currents is one of the basic principles underlying the rapidly expanding field of spintronics. However, no direct demonstration of the phenomenon exists. We report a novel measurement in which a nonequilibrium spin population is created by a pointlike injection of current from a ferromagnet across a tunnel barrier into a one-dimensional spin channel and detected differentially by a pair of ferromagnetic electrodes placed symmetrically about the injection point. We demonstrate that the spin current is strictly isotropic about the injection point and, therefore, completely decoupled from the unidirectional charge current.

Giant fluctuations of superconducting order parameter in ferromagnet-superconductor single-electron transistors.

Spin dependent transport in a ferromagnet-superconductor single-electron transistor is studied theoretically taking into account spin accumulation, spin relaxation, gap suppression, and charging effects. A strong dependence of the gap on the magnetic state of the electrodes is found, which gives rise to a magnetoresistance of up to 100%. We predict that fluctuations of the spin accumulation can play such an important role as to cause the island to fluctuate between the superconducting and normal states. Furthermore, the device exhibits a nearly complete gate-controlled spin-valve effect.

Coulomb blockade and coherent single-cooper-pair tunneling in single Josephson junctions.

We have measured the current-voltage characteristics of small-capacitance single Josephson junctions at low temperatures ( T< or =0.04 K), where the strength of the coupling between the single junction and the electromagnetic environment was controlled with one-dimensional arrays of dc SQUIDs. We have clearly observed Coulomb blockade of Cooper-pair tunneling and even a region of negative differential resistance, when the zero-bias resistance of the SQUID arrays is much higher than the quantum resistance h/e(2) approximately 26 kOmega. The negative differential resistance is evidence of coherent single-Cooper-pair tunneling in the single Josephson junction.

Cloning, expression, sequence determination, and chromosome localization of the mouse complement C3a anaphylatoxin receptor gene.

The complement C3a anaphylatoxin receptor (C3aR) is a seven-transmembrane G-protein coupled chemoattractant receptor that on binding the C3a peptide ligand mediates numerous cellular responses, including histamine release from mast cells. smooth muscle contraction, and the directed migration of eosinophils. To delineate the murine C3aR coding sequence, gene structure, 5'-flanking region, and chromosome location, cDNA and genomic clones encoding the mouse C3a receptor were isolated, characterized, and used in fluorescence in situ hybridization experiments. The results from this study indicate that the murine C3a receptor structural gene is a single copy gene of approximately 8 kb comprised of 2 exons which are separated by a large intervening intron of 4724 bp. The first exon encodes 97 bp of 5'-untranslated sequence. Exon 2 encodes the remaining 8 bp of 5'-untranslated sequence and the entire coding and 3'-untranslated sequences. This genomic organization is typical of most other chemoattractant receptor genes in that the entire coding sequence is contained on a single exon. The human and mouse C3a receptor genes were localized to syntenic chromosomal bands 12q13.2-3 and 6F1, respectively. No other seven-transmembrane receptor genes, to date, have been localized to these chromosomal regions. Primer extension experiments using mouse macrophage RNA indicated a single transcriptional initiation site. Sequence analysis 5' of the transcriptional site indicated a TATA-less promoter with possible cis-acting motifs that may regulate C3a receptor gene expression. These included the recognition sequence for the nuclear transcription factor SP1 and the phorbol ester response sequence which binds the Fos/Jun heteromeric transcription factor AP1.

Expression of the receptors for the C5a anaphylatoxin, interleukin-8 and FMLP by human astrocytes and microglia.

The expression of chemotactic receptors in the central nervous system is largely unexplored. In this study, we examined human astrocytes and microglia as well as the conditionally immortalized human astrocyte cell line HSC2 for expression of the C5a-anaphylatoxin receptor (C5aR), the interleukin-8 receptor (IL-8R) and the f-Met-Leu-Phe receptor (FMLPR). Using flow cytometry, indirect immunofluorescence and RT-PCR analysis, we demonstrated that astrocytes, microglia and HSC2 cells contain specific RNA and express surface protein for all three chemotactic receptors. These are the first studies to demonstrate definitively the expression of these chemotactic receptors astrocytes and microglia, thereby expanding the types of cells known to express chemotactic receptors. Moreover, these data suggest that these chemotactic receptors may play an important role in mediating the inflammatory response and perhaps other yet undescribed biological phenomena in the central nervous system.

N-formylpeptide and complement C5a receptors are expressed in liver cells and mediate hepatic acute phase gene regulation.

Although the classical chemotactic receptor for complement anaphylatoxin C5a has been associated with polymorphonuclear and mononuclear phagocytes, several recent studies have indicated that this receptor is expressed on nonmyeloid cells including human endothelial cells, vascular smooth muscle cells, bronchial and alveolar epithelial cells, hepatocytes, and in the human hepatoma cell line HepG2. In this study, we examined the possibility that other members of the chemotactic receptor family are expressed in HepG2 cells and human liver, and the possibility that such receptors mediate changes in acute phase gene expression in HepG2 cells. Using polymerase chain reaction (PCR) amplification of HepG2 mRNA with primers based on highly conserved regions of the chemotactic subgroup of the G protein-coupled receptor family, we identified a PCR fragment from the formyl-methionyl-leucyl-phenylalanine (FMLP) receptor, as well as one from the C5a receptor. Immunostaining with antipeptide antisera to FMLPR confirmed the presence of this receptor in HepG2 cells. Receptor binding studies showed specific saturable binding of a radioiodinated FMLP analogue to HepG2 cells (Kd approximately 2.47 nM; R approximately 6 x 10(3) plasma membrane receptors per cell). In situ hybridization analysis showed the presence of FMLPR mRNA in parenchymal cells of the human liver in vivo. Both C5a and FMLP mediated concentration- and time-dependent changes in synthesis of acute phase proteins in HepG2 cells including increases in complement C3, factor B, and alpha 1-antichymotrypsin, as well as concomitant decreases in albumin and transferrin synthesis. The effects of C5a and FMLP on the synthesis of these acute phase proteins was evident at concentrations as low as 1 nM, and they were specifically blocked by antipeptide antisera for the corresponding receptor. In contrast to the effect of other mediators of hepatic acute phase gene regulation, such as interleukin 6, the effects of C5a and FMLP were reversed by increased concentrations well above the saturation point of the respective receptor. These results suggest that acute phase gene regulation by C5a and FMLP is desensitized at high concentrations, a property that is unique among the several known mechanisms for hepatic acute phase gene regulation.

Inherited human complement C5 deficiency. Nonsense mutations in exons 1 (Gln1 to Stop) and 36 (Arg1458 to Stop) and compound heterozygosity in three African-American families.

Hereditary C5 deficiency has been reported in several families of different ethnic backgrounds and from different geographic regions, but the molecular genetic defect causing C5 deficiency has not been delineated in any of them. To examine the molecular basis of C5 deficiency in the African-American population, the exons and intron/exon boundaries of the C5 structural genes from three C5-deficient (C5D) African-American families were sequenced, revealing two nonsense mutations. The nonsense mutations are located in exon 1 (C84AG to TAG) in two of the C5D families (Rhode Island and North Carolina) and in exon 36 (C4521GA to TGA) in the third C5D family (New York). The exon 1 and 36 mutations are contained in codons that encode the first amino acid of the C5 beta-chain (Gln1 to Stop) and residue 1458 in the alpha-chain (Arg1458 to Stop), respectively. Allele-specific PCR and sequence analyses demonstrated that the exon 1 mutation is present in only one of the C5 null genes in both the Rhode Island and North Carolina families, and the exon 36 mutation is contained in only one C5 null gene in the New York family. Neither of the nonsense mutations was found in the European or Caucasian-American C5D individuals examined. Collectively, these data indicate that: 1) C5 deficiency is caused by several different molecular genetic defects, 2) C5 deficiency in the African-American population can be explained in part by two distinct nonsense mutations in exons 1 and 36, and 3) compound heterozygosity exists in all of the reported African-American C5D families.

Cellular expression of the C5a anaphylatoxin receptor (C5aR): demonstration of C5aR on nonmyeloid cells of the liver and lung.

The small-complement C5 activation fragment, C5a, is a potent phlogistic molecule that, on binding to the C5a Receptor (C5aR), mediates contraction of smooth muscle, enhances vascular permeability, and promotes leukocyte functions such as directed chemotaxis, degranulation, mediator release, and production of superoxide anions. Although C5aR expression has traditionally been thought to be limited primarily to myeloid blood cells, including neutrophils, monocytes, macrophages, and eosinophils, we report here that C5aR is expressed by liver and lung cells as well as by cells in several other tissues. By Northern blot analysis, it was determined that mouse liver, baboon liver, human liver, and the human hepatoma-derived cell line HepG2 express a normal size (2.3 kb) C5aR mRNA; in HepG2 cells, the quantity of C5aR mRNA was comparable to that contained in dbcAMP-differentiated U937 cells. HepG2 cells were demonstrated to express the C5aR on their cell surface by flow cytometric and immunofluorescence analyses as well as by 125I-C5a binding assays. The binding data indicated that HepG2 cells express a single class of C5aR with a Kd of 1.18 nM and approximately 28,000 receptors per cell. In vivo expression of C5aR in human liver cells was demonstrated by in situ hybridization and immunohistochemistry analyses. Northern blot analysis of murine and baboon organs shows that, in addition to the liver, other tissues express C5aR mRNA in significant quantities, including the spleen, lung, heart, kidney, and intestine. Moreover, mice treated with LPS show a large increase in C5aR mRNA in all these tissues except the intestine. Immunostaining of human lung tissue demonstrated that bronchial and alveolar epithelial cells, as well as vascular smooth muscle and endothelial cells, also express the C5aR. Collectively, these data indicate that the C5aR is expressed in several different types of cells in liver and lung, and in yet undetermined cell types in spleen, heart, intestine, and kidney. Furthermore, these data suggest that the C5a anaphylatoxin mediates previously unrecognized functions by binding to tissue cells that express the C5aR.

Structure, 5'-flanking sequence, and chromosome location of the human N-formyl peptide receptor gene. A single-copy gene comprised of two exons on chromosome 19q.13.3 that yields two distinct transcripts by alternative polyadenylation.

The N-formyl peptide chemoattractant receptor (fMLF-R) is a cell-surface, G-protein-coupled glycoprotein that mediates the directed locomotion of neutrophils upon binding N-formylated peptides. The fMLF-R is encoded primarily by a 1.6-kb mRNA in differentiated HL-60 and U937 cells, although larger less abundant transcripts are present. To study the origin of different fMLF-R transcripts, the genetic linkage of chemotactic receptor genes, and the regulation of fMLF-R gene expression, we determined the copy number, chromosomal location, structural organization, and 5'-flanking sequence of the human fMLF-R gene. BamHI restriction fragments derived from a human fMLF-R genomic cosmid clone were isolated, subcloned, and sequenced. These data indicate that the fMLF-R structural gene is approximately 7.5 kb in length and is comprised of two exons separated by an approximately 5.0-kb intron. The first exon encodes 66 bp of the 5'-untranslated sequence, while exon 2 encodes the coding and 3'-untranslated sequences. The genomic organization of the fMLF-R gene is similar to that of the adrenergic beta-1 and beta-2 G-protein-coupled receptor genes in that the coding sequence is contained in a single exon. The different 3'-untranslated sequences observed in fMLF-R cDNA clones are contiguous in the genomic structure, thereby indicating that these clones are derived in part by alternative polyadenylation. Southern blot analysis using human X hamster somatic cell hybrids and in situ hybridization indicated that the h-fMLF-R gene is located on chromosome 19q13.3. Primer extension experiments using dbcAMP-differentiated U937 RNA indicated a single transcriptional initiation site. Sequence analysis 5' of the transcriptional initiation site indicated possible cis-acting motifs that may regulate fMLF-R gene expression. These included AP-1 and CK-2 consensus sequences that bind nuclear factors of the Fos/Jun family and NF-GMb, respectively.

Structural aspects of the human C5 gene. Intron/exon organization, 5'-flanking region features, and characterization of two truncated cDNA clones.

Human C5 cDNA fragments were used to identify five overlapping cosmid clones that spanned the entire C5 gene. Partial sequencing and Southern analysis of the clones were performed to identify intron/exon boundaries and to map intron size. The human C5 gene is 79 kilobases in length and is comprised of 41 exons. Comparison of C5 with the homologous family members C3 and C4 revealed striking similarities in exon size and number. Less, although significant similarities were also observed with the family member alpha 2-macroglobulin. The transcriptional start site for the C5 gene was observed as a doublet at positions 29 and 28 nucleotides upstream of the ATG start codon. The 5'-flanking region of the gene contains sequences homologous with several known responsive elements, including interferon, interleukin-6, glucocorticoid, estrogen, NF-kappa B, and HNF-1. Two previously identified truncated cDNAs, pHC5A and pHC5B, contain 21 and 16 exons, respectively. The last exon in pHC5A, designated exon 21a, is a product of alternative splicing and is not present in the major full-length transcript. Truncation of pHC5A is the result of an alternative polyadenylation signal located in exon 21a. In pHC5B, exon 16 is extended on the 3' end by additional flanking genomic sequence that also contains an alternative polyadenylation signal.

Structure of the murine fifth complement component (C5) gene. A large, highly interrupted gene with a variant donor splice site and organizational homology with the third and fourth complement component genes.

To understand fifth complement component (C5) gene regulation, splicing, and C5 protein deficiency at the molecular level, the organization of the murine C5 gene was determined. The C5 structural gene is present as a single copy in the mouse genome as demonstrated by Southern blot analysis. Accordingly, three cosmid clones were isolated from a genomic library that was prepared from mouse strain B10.D2/nSnJ. These clones overlapped and contained the structural gene encoding the complete C5 alpha-chain and 90% of the beta-chain. The 5'-flanking region of the C5 gene was obtained from a clone isolated from a genomic lambda-MOPC-41 library. Unique restriction fragments were prepared from the genomic clones and subcloned, and the exons were sequenced. All introns were sized by sequencing or Southern analysis. The C5 structural gene was found to be a highly interrupted gene of approximately 78 kilobases containing 42 exons and 41 introns. The exons ranged in length from 58 to 247 base pairs, with an average length of 131 base pairs. The introns ranged in size from 100 base pairs to 4 kilobases with an average length of 1.5 kilobases. The C5 alpha-chain was encoded by 49 kilobases containing 26 exons; the beta-chain was encoded by 29 kilobases containing 16 exons. The C5a coding sequence was split between two exons. All intron/exon junctions followed the normal consensus rule except at intron 35 in which the 5'-donor GT was substituted by GC. The 2-base-pair gene deletion and HindIII and PvuII restriction fragment length polymorphisms associated with murine C5 deficiency were localized to exon 7, exon 16, and intron 20, respectively. Comparison of the intron-exon junctions of the murine C5, human C3, and mouse C4 genes indicated that these genes are nearly identical in structural organization. However, the rat alpha 2-macroglobulin gene showed only moderate genomic organizational similarity to the murine C5 gene. A major and a minor transcriptional initiation site in the C5 gene were identified by primer extensions and confirmed by RNase protection assays. Sequence analysis of the 5'-flanking region (760 base pairs) revealed a TATA-like and CAAT box upstream of the major transcriptional initiation site at positions -274 and -303, respectively, suggesting an atypical promoter. The 5'-flanking region also contained sequences identical with several cis-acting motifs known to bind the liver-specific nuclear protein LF-A1 and the nuclear protein NF-kappa B.(ABSTRACT TRUNCATED AT 250 WORDS)