Monitoring Heterogeneously Catalyzed Hydrogenation Reactions at Elevated Pressures Using In-Line Flow NMR.

We present a novel setup that can be used for the in-line monitoring of solid-catalyzed gas-liquid reactions. The method combines the high sensitivity and resolution of a stripline NMR detector with a microfluidic network that can withstand elevated pressures. In our setup we dissolve hydrogen gas in the solvent, then flow it with the added substrate through a catalyst cartridge, and finally flow the reaction mixture directly through the stripline NMR detector. The method is quantitative and can be used to determine the solubility of hydrogen gas in liquids; it allows in-line monitoring of hydrogenation reactions and can be used to determine the reaction kinetics of these reactions. In this work, as proof of concept we demonstrate the optimization of the Pd-catalyzed hydrogenation reactions of styrene, phenylacetylene, cyclohexene, and hex-5-en-2-one in a microfluidic context.

Pretransplant NPM1 MRD levels predict outcome after allogeneic hematopoietic stem cell transplantation in patients with acute myeloid leukemia.

The objective was to evaluate the prognostic impact of pre-transplant minimal residual disease (MRD) as determined by real-time quantitative polymerase chain reaction in 67 adult NPM1-mutated acute myeloid leukemia patients receiving allogeneic hematopoietic stem cell transplantation (HSCT). Twenty-eight of the 67 patients had a FLT3-ITD (42%). Median age at transplantation was 54.7 years, median follow-up for survival from time of allografting was 4.9 years. At transplantation, 31 patients were in first, 20 in second complete remission (CR) and 16 had refractory disease (RD). Pre-transplant NPM1 MRD levels were measured in 39 CR patients. Overall survival (OS) for patients transplanted in CR was significantly longer as compared to patients with RD (P=0.004), irrespective of whether the patients were transplanted in first or second CR (P=0.74). There was a highly significant difference in OS after allogeneic HSCT between pre-transplant MRD-positive and MRD-negative patients (estimated 5-year OS rates of 40 vs 89%; P=0.007). Multivariable analyses on time to relapse and OS revealed pre-transplant NPM1 MRD levels >1% as an independent prognostic factor for poor survival after allogeneic HSCT, whereas FLT3-ITD had no impact. Notably, outcome of patients with pre-transplant NPM1 MRD positivity >1% was as poor as that of patients transplanted with RD.

Spatially resolved spectroscopy using tapered stripline NMR.

Magnetic field B0 gradients are essential in modern Nuclear Magnetic Resonance spectroscopy and imaging. Although RF/B1 gradients can be used to fulfill a similar role, this is not used in common practice because of practical limitations in the design of B1 gradient coils. Here we present a new method to create B1 gradients using stripline RF coils. The conductor-width of a stripline NMR chip and the strength of its radiofrequency field are correlated, so a stripline chip can be tapered to produce any arbitrary shaped B1 field gradient. Here we show the characterization of this tapered stripline configuration and demonstrate three applications: magnetic resonance imaging on samples with nL-µL volumes, reaction monitoring of fast chemical reactions (10(-2)-10(1)s) and the compensation of B0 field gradients to obtain high-resolution spectra in inhomogeneous magnetic fields.

High resolution triple resonance micro magic angle spinning NMR spectroscopy of nanoliter sample volumes.

To be able to study mass-limited samples and small single crystals, a triple resonance micro-magic angle spinning (µMAS) probehead for the application of high-resolution solid-state NMR of nanoliter samples was developed. Due to its excellent rf performance this allows us to explore the limits of proton NMR resolution in strongly coupled solids. Using homonuclear decoupling we obtain unprecedented (1)H linewidths for a single crystal of glycine (Δν(CH2) = 0.14 ppm) at high field (20 T) in a directly detected spectrum. The triple channel design allowed the recording of high-resolution µMAS (13)C-(15)N correlations of [U-(13)C-(15)N] arginine HCl and shows that the superior (1)H resolution opens the way for high-sensitivity inverse detection of heteronuclei even at moderate spinning speeds and rf-fields. Efficient decoupling leads to long coherence times which can be exploited in many correlation experiments.

Craniofrontonasal syndrome in a male due to chromosomal mosaicism involving EFNB1: further insights into a genetic paradox.

Craniofrontonasal syndrome (CFNS) is an X-linked disorder caused by inactivating mutations in the gene for ephrin-B1 (EFNB1). Paradoxically it shows a more severe phenotype in females than in males. As a result of X inactivation cell populations with and without EFNB1 expression are found in EFNB1+/- females. This is thought to initiate a process termed cellular interference which may be responsible for the phenotype in females. We present a boy with severe clinical features of CFNS. In ∼42% of his blood cells we found a supernumerary ring X chromosome containing EFNB1 but lacking XIST. Mosaicism for cell populations with different levels of EFNB1 expression can explain the severe phenotype of this patient. In vitro experiments in Xenopus tissue showed that cells overexpress ephrinB1 cluster and sort out from wild-type cells. Our report provides further evidence that cellular interference contributes to the paradoxical inheritance pattern of CFNS.

Silver-Russell syndrome due to maternal uniparental disomy 7 and a familial reciprocal translocation t(7;13).

Silver-Russell syndrome (SRS) is a genetically heterogeneous disorder characterized by intrauterine and postnatal growth retardation, typical facial features and a spectrum of additional features including body and limb asymmetry and clinodactyly. Maternal uniparental disomy for chromosome 7 (upd(7)mat) was shown to occur in 5-10% of patients with SRS. Maternal UPD7 is clinically often associated with mild SRS. Parents of an affected child are given a negligible recurrence risk as all reported cases with upd(7)mat have been sporadic so far. In general, chromosomal rearrangements-like translocations increase the likelihood of uniparental disomy (UPD) for the chromosomes involved. However, SRS as the result of a upd(7)mat in association with an inherited chromosomal translocation involving chromosome 7 has only been reported once before. Here, we describe the second case of SRS with upd(7)mat due to a familial reciprocal translocation t(7;13). This emphasizes the importance of chromosome analysis in SRS patients with upd(7)mat to rule out chromosomal rearrangements despite their rare occurrence as they are of great relevance for genetic counseling of SRS families.

Pseudoautosomal inheritance of Léri-Weill syndrome: what does it mean?

The short stature homeobox (SHOX) gene is located in the pseudoautosomal region 1 of both sex chromosomes. Haploinsufficiency of SHOX leads to different phenotypes ranging from isolated short stature to Léri-Weill syndrome characterized by short stature, mesomelia and Madelung deformity. We describe a family with a SHOX deletion originally located on the Y chromosome and transmitted from father to daughter by crossover during meiosis. The male index patient presented with short stature, mesomelia and mild Madelung deformity. His father had a normal height but slightly disproportionate short legs. The sister of the index patient presented with marked Madelung deformity and normal height. A deletion of the SHOX gene was identified in the male index patient, his father and his sister. Metaphase fluorescence in situ hybridization (FISH) analyses showed a deletion of the SHOX gene on the Y chromosomes of the index patient and his father, and on the X chromosome of his sister, indicating that a meiotic crossover of the SHOX gene region between the X and Y chromosomes had occurred. The pseudoautosomal region 1 is a known recombination 'hot spot' in male meiosis. Published genetic maps indicate high recombination frequency of ∼40% for SHOX in male meiosis leading to pseudoautosomal inheritance.

Full quadrupolar tensor determination by NMR using a micro-crystal spinning at the magic angle.

An implementation of rotor-synchronised Magic Angle Spinning (MAS) NMR is presented to determine the quadrupolar coupling tensor values from a single crystal study for half-integer quadrupolar nuclei. Using a microcoil based probehead for studying micro crystals with superior sensitivity, we successfully determine the full quadrupolar tensor of (23)Na using a micro crystal of dimensions 210 x 210 x 700 mum of NaNO(3) as a model system. A two step simulation procedure is used to obtain the orientation of the quadrupolar tensor information from the experimental spectra and is verified by XRD analysis.

Optimization of stripline-based microfluidic chips for high-resolution NMR.

We here report on the optimization, fabrication and experimental characterization of a stripline-based microfluidic NMR probe, realized in a silicon substrate. The stripline geometry was modelled in respect of rf-homogeneity, sensitivity and spectral resolution. Using these models, optimal dimensional ratios were found, which hold for every sample size. Based on the optimized parameters, a simple integrated stripline-based microfluidic chip was realized. The fabrication of this chip is described in detail. We achieved a sensitivity of 0.47 nmol/square root(Hz) and a resolution of 0.7 Hz. The rf-homogeneity (A(810 degrees)/A(90 degrees)) was 76% and was proved to be suitable for 2D-NMR analysis of glucose.

High-resolution liquid- and solid-state nuclear magnetic resonance of nanoliter sample volumes using microcoil detectors.

The predominant means to detect nuclear magnetic resonance (NMR) is to monitor the voltage induced in a radiofrequency coil by the precessing magnetization. To address the sensitivity of NMR for mass-limited samples it is worthwhile to miniaturize this detector coil. Although making smaller coils seems a trivial step, the challenges in the design of microcoil probeheads are to get the highest possible sensitivity while maintaining high resolution and keeping the versatility to apply all known NMR experiments. This means that the coils have to be optimized for a given sample geometry, circuit losses should be avoided, susceptibility broadening due to probe materials has to be minimized, and finally the B(1)-fields generated by the rf coils should be homogeneous over the sample volume. This contribution compares three designs that have been miniaturized for NMR detection: solenoid coils, flat helical coils, and the novel stripline and microslot designs. So far most emphasis in microcoil research was in liquid-state NMR. This contribution gives an overview of the state of the art of microcoil solid-state NMR by reviewing literature data and showing the latest results in the development of static and micro magic angle spinning (microMAS) solenoid-based probeheads. Besides their mass sensitivity, microcoils can also generate tremendously high rf fields which are very useful in various solid-state NMR experiments. The benefits of the stripline geometry for studying thin films are shown. This geometry also proves to be a superior solution for microfluidic NMR implementations in terms of sensitivity and resolution.

Stripline probes for nuclear magnetic resonance.

A novel route towards chip integrated NMR analysis is evaluated. The basic element in the design is a stripline RF 'coil' which can be defined in a single layer lithographic process and which is fully scalable to smaller dimensions. The sensitivity of such a planar structure can be superior to that of a conventional 3D helix. The basic properties, such as RF field strength, homogeneity and susceptibility broadening are discussed in detail. Secondary effects related to the thermal characteristics are discussed in simplified models. Preliminary NMR tests of basic solid and liquid samples measured at 600 MHz confirm the central findings of the design study. It is concluded that the stripline structure can be a valuable addition to the NMR toolbox; it combines high sensitivity with low susceptibility broadening and high power handling capabilities in a simple scalable design.

Towards nuclear magnetic resonance micro-spectroscopy and micro-imaging.

The first successful experiments demonstrating Nuclear Magnetic Resonance (NMR) were a spin-off from the development of electromagnetic technology and its introduction into civilian life in the late forties. It was soon discovered that NMR spectra held chemically relevant information making it useful as an analytical tool. By introducing a new way of detection, moving away from continuous wave spectroscopy, Fourier Transform NMR helped to overcome sensitivity problems and subsequently opened the way for multi-dimensional spectroscopy. As a result NMR has developed into one of the most powerful analysis techniques with widespread applications. Still sensitivity is a limiting factor in the applicability of NMR. Therefore we witness a renaissance of technique development in magnetic resonance striving to improve its receptiveness. This tutorial review introduces the efforts currently made in miniaturizing inductive detection by designing optimal radio-frequency microcoils. A second approach is to introduce a new way of detecting magnetic resonance signals by means of very sensitive micromechanical force detectors. This shows that the detection limits in terms of absolute sensitivity or imaging resolution are still open to significant improvements.

Implementing solenoid microcoils for wide-line solid-state NMR.

Solid-state nuclear magnetic resonance (NMR) probeheads using solenoid microcoils with an inner diameter of 300-400 microm are developed for the study of mass-limited solid samples. Some attention is paid to the mechanical ruggedness of the probes allowing sample changing. The performance, in terms of sensitivity and RF-characteristics, of these probeheads is studied for (1)H, (31)P, and (27)Al in different model compounds in view of the feasibility of specific applications. The results show that the sensitivity is high enough to detect approximately 10(14) spins/sqrt Hz with a signal-to-noise ratio of 1 in a single scan. A specific advantage of microcoils for solid-state NMR applications is that they can generate extremely high RF-fields if implemented in appropriate circuits. Using RF-powers in the hundreds of Watts range, RF-fields well in excess of 1MHz can be made. This allows the excitation of spectra of nuclei whose resonance lines are dispersed of several megahertz. This is particularly useful for quadrupolar nuclei experiencing large quadrupolar interactions as is demonstrated for (27)Al in single crystal and powdered minerals.

Disruption of the RanBP17/Hox11L2 region by recombination with the TCRdelta locus in acute lymphoblastic leukemias with t(5;14)(q34;q11).

The t(5;14)(q33-34;q11) translocation constitutes a recurrent rearrangement in acute lymphoblastic leukemia involving the T cell receptor (TCR) delta locus on chromosome 14. Breakpoint sequences of the derivative chromosome 5 were isolated by application of a ligation-mediated PCR technique using TCR delta-specific primers to amplify genomic DNA from the leukemic cells of a patient with t(5;14). Through exon trap analysis, we identified various putative exons of the chromosome 5 target gene of the translocation; compilation of sequence information of trapped exons and available expressed sequence tags (ESTs) from the GenBank database allowed us to assemble 1.2 kb of the cDNA. Full-length cDNAs were isolated from a human testis cDNA library and sequence analysis predicted a putative Ran binding protein, a novel member of the importin-beta superfamily of nuclear transport receptors, called RanBP17. The t(5;14) breakpoint maps to the 3' coding region of the gene. The breakpoint of a second t(5;14) positive patient was mapped about 8 kb downstream of the most 3' RanBP17 exon and 2 kb upstream of the first exon of the orphan homeobox gene, Hox11L2. In both cases TCR delta enhancer sequences are juxtaposed downstream of the truncated or intact RanBP17 gene, respectively on the derivative chromosome.