Use of a small particle solid‐core packing for improved efficiency and rapid measurement of sirolimus and everolimus by LC‐MS/MS

Measurement of whole blood sirolimus and everolimus is required in order to optimize patient treatment following solid organ transplant. Assay by LC‐MS/MS is increasingly preferred; however efficient use of the instrument and short turnaround times are crucial. Use of a 1.6 µm solid‐core packing HPLC column (Cortecs) gave significant increases in efficiency, sensitivity and throughput compared with an existing method, following simple protein precipitation of small‐volume (20 μL) whole blood samples. Sirolimus, everolimus and the stable isotopic internal standard (¹³C₂D₄ – everolimus) eluted at around 0.8 min, and total analytical run time was 2.2 min, saving almost 4 min per sample compared with an existing method. Within‐assay imprecision (CV) was 3.3–8.5%, and between‐assay imprecision was 2.2–10.8%. Retrospective assay of external quality assurance samples and comparison of patient samples assayed in parallel showed only small differences (between +6.8 and ?1.9%) in results using the Cortecs column when compared with the existing method. No significant interferences or ion suppression were observed. Copyright © 2015 John Wiley & Sons, Ltd.     

Newton's method on Riemannian manifolds and a geometric model for the human spine

To study a geometric model of the human spine we are led tofinding a constrained minimum of a real valued function definedon a product of special orthogonal groups. To take advantgeof its Lie group structure we consider Newton's method on thismanifold. Comparisons between measured spines and computed spinesshow the pertinence of this approach.     

Use of labelled nitrogen to measure gross and net rates of mineralization and microbial activity in permanent pastures following fertilizer applications at different time intervals

Measurements of some of the main internal N-cycling processes in soil were obtained by labelling the inorganic N pool with the stable isotope of nitrogen ((15)N). The (15)N mean pool dilution technique, combined with other field measurements, enabled gross and net N-mineralization rates to be resolved in grassland soils, which had previously either received fertilizer N (F), or had remained unfertilized (U) for many years. The two soils were subdivided into plots that received N at different time intervals (over 3 weeks), prior to (15)N measurements being made. By this novel approach, possible 'priming' effects over time were investigated to try to overcome some of the temporal problems of isotopic labelling of soil N (native plus fertilizer) and to identify possible changes in a range of primary N-transformation processes. The results suggested that an overall stimulation of microbially mediated processes occurred with all N treatments, but there were inconsistencies associated with the release of N, both in the timing and the degree to which different processes responded to the application of fertilizer N. The rates of these processes were, however, within the range of previously reported data and the (15)N measurements were not adversely affected by the differences in N pools created by the treatments. Thus, the mean pool dilution technique was shown to be applicable to agricultural soils, under conditions relevant to grass swards receiving fertilizer. For example, between the U and F treatments, the size of inorganic N pools increased by five-fold and gross rates of mineralization reached 3.5 and 4.8 microg N g(-1) (dry soil) d(-1), respectively, but did not vary greatly with the timing of N applications. A correlation (r(2) = 0.57) was found between soil respiration (which is relatively simple to measure) and net mineralization (which is more time consuming), suggesting that the former might be used as an indicator of the latter. Although this relationship was stronger in previously unfertilized soils, the similarities found with fertilized soils suggest that this approach could be used to obtain information of wider agronomic value and would, therefore, warrant further work under a range of soil conditions.     

Manpower Planning in the National Health Service

Journal of the Operational Research Society -     

Profiling of generic anti-phosphopeptide antibodies and kinases with peptide microarrays using radioactive and fluorescence-based assays

Kinases represent one of the largest enzyme families and key regulatory proteins in the cell. Only a small subset of these enzymes has been characterised so far. We have prepared different types of phosphopeptide and peptide microarrays displaying peptides deduced from annotated human phosphorylation sites and cytoplasmic domains of all annotated human membrane proteins. This approach was enabled by fully-automated high throughput micro-scale synthesis of peptides by the SPOT technology combined with chemo-selective immobilisation on modified glass slides. The phosphopeptide microarrays displaying 2923 peptides in total have been used for the characterisation of commercially available generic anti-phosphopeptide antibodies. This enabled us to detect Abl kinase activity on a microarray with anti-phosphotyrosine antibodies yielding results comparable to those obtained from a radioactive assay. More than 13 000 peptides deposited on six glass slides were used to profile casein kinase 2 (CK2) using a radioactive assay, since no generic antibody for the reliable detection of serine or threonine phosphorylation could be identified. All previously identified substrates were detected in the microarray experiment. In order to confirm whether substrates on the microarray are substrates in solution phase assays, more than 700 peptides were synthesised and tested with CK2 in a solution phase assay. All substrates identified in the solution phase assay were also detected on the microarray.     

A mixed ladderane/n-alkyl glycerol diether membrane lipid in an anaerobic ammonium-oxidizing bacterium

A novel glycerol diether containing ladderane and tetradecyl moieties has been identified in an anaerobic ammonium-oxidizing bacterium by GC/MS and high-field NMR spectroscopy.     

Field-induced thermal metal-to-insulator transition in underdoped La(2-x)Sr(x)CuO(4+delta)

The transport of heat and charge in cuprates was measured in single crystals of La(2-x)Sr(x)CuO(4+delta) (LSCO) across the doping phase diagram at low temperatures. In underdoped LSCO, the thermal conductivity is found to decrease with increasing magnetic field in the T-->0 limit, in striking contrast to the increase observed in all superconductors, including cuprates at higher doping. In heavily underdoped LSCO, where superconductivity can be entirely suppressed with an applied magnetic field, we show that a novel thermal metal-to-insulator transition takes place upon going from the superconducting state to the field-induced normal state.