Spectral imaging for precise surgical intervention in Hirschsprung's Disease

We used advanced spectral imaging for intrasurgical decision making in a preclinical study, on a mouse model of Hirschsprung's Disease. Our imaging device sampled areas from normal and abnormal (aganglionic) colon in these animals. Spectral segmentation and classification of the resulting images showed a clear distinction between the normal and aganglionic regions, as confirmed by pathological analysis and use of mutant mice. We developed a simple algorithm that could distinguish normal from aganglionic colon with high spatial resolution and reproducibility, and the following statistics: sensitivity = 97%, specificity = 94%, positive predictive value = 92%, negative predictive value = 98%. These studies showed translational proof of concept that spectral imaging could be used during operations, in real time, to help surgeons precisely distinguish normal from abnormal tissue without requiring traditional biopsy. (© 2008 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

Raman-based dynamic feeding strategies using real-time glucose concentration monitoring system during adalimumab producing CHO cell cultivation

The use of Process Analytical Technology tools coupled with chemometrics has been shown great potential for better understanding and control of mammalian cell cultivations through real-time process monitoring. In-line Raman spectroscopy was utilized to determine the glucose concentration of the complex bioreactor culture medium ensuring real-time information for our process control system. This work demonstrates a simple and fast method to achieve a robust partial least squares calibration model under laboratory conditions in an early phase of the development utilizing shake flask and bioreactor cultures. Two types of dynamic feeding strategies were accomplished where the multi-component feed medium additions were controlled manually and automatically based on the Raman monitored glucose concentration. The impact of these dynamic feedings was also investigated and compared to the traditional bolus feeding strategy on cellular metabolism, cell growth, productivity, and binding activity of the antibody product. Both manual and automated dynamic feeding strategies were successfully applied to maintain the glucose concentration within a narrower and lower concentration range. Thus, besides glucose, the glutamate was also limited at low level leading to reduced production of inhibitory metabolites, such as lactate and ammonia. Consequently, these feeding control strategies enabled to provide beneficial cultivation environment for the cells. In both experiments, higher cell growth and prolonged viable cell cultivation were achieved which in turn led to increased antibody product concentration compared to the reference bolus feeding cultivation.     

Exploring context dependency in eco‐evolutionary patterns with the stick insect Timema cristinae

Rapid evolution can influence the ecology of populations, communities, and ecosystems, but the importance of evolution for ecological dynamics remains unclear, largely because the contexts in which evolution is powerful are poorly resolved. Here, we carry out a large observational study to test hypotheses about context dependency of eco‐evolutionary patterns previously identified on the stick insect Timema cristinae. Experiments and observations conducted in 2011 and 2012 documented predator‐mediated negative effects of camouflage maladaptation (i.e., evolutionary dynamics) on: (a) T. cristinae abundance and, (b) species richness and abundance of other arthropods. Here we show that camouflage maladaptation does not correlate with T. cristinae abundance and, instead, is associated with increased abundance and species richness of cohabitating arthropods. We furthermore find that plants with high levels of Timema maladaptation tend to have higher foliar nitrogen, that is, higher nutritional value, and more positive mass‐abundance slopes in the coexisting arthropod communities. We propose explanations for the observed contrasting results, such as negative density‐ and frequency‐dependent selection, feedbacks between herbivore abundance and plant nutritional quality, and common effects of predation pressure on selection and prey abundance. Our results demonstrate the utility of observational studies to assess the context dependency of eco‐evolutionary dynamics patterns and provide testable hypotheses for future work.     

Nucleosides,XLIII1) Synthesis and Properties of 04-Alkylthymidines

Abstract

Various 0⁴-alkylthymidines 14–20 have been synthesized by two different methods. 0⁴-Alkylation takes place with 3′,5′-di-0-acetyl- (2) and 3′,5′-di-0-benzoylthymidine (3) respectively in a silver ion catalysed reaction with alkyl halides, whereas the azolide approach makes use of a nucleophilic displacement of the appropriate intermediate by alkoxides and subsequent deacylation to the free nucleosides. Structural proofs are based on elemental analyses, UV- and ¹H-NMR-spectra.     

Recent Progress in Oligoribonucleotide Synthesis

Abstract

The usefulness of the p-nitrophenylethylsulfonyl (NPES) group for 2′-OH protection in oligoribonucleotide synthesis is further investigated. The difficulties of uridine protection are discussed and the p-cyanophenylethyl (CPE) group introduced as a versatile new 0⁴-blocking group.     

Improved growth media and culture techniques for genetic analysis and assessment of biomass utilization by Caldicellulosiruptor bescii

Methods for efficient growth and manipulation of relatively uncharacterized bacteria facilitate their study and are essential for genetic manipulation. We report new growth media and culture techniques for Caldicellulosiruptor bescii, the most thermophilic cellulolytic bacterium known. A low osmolarity defined growth medium (LOD) was developed that avoids problems associated with precipitates that form in previously reported media allowing the monitoring of culture density by optical density at 680 nm (OD₆₈₀) and more efficient DNA transformation by electroporation. This is a defined minimal medium and does not support growth when a carbon source is omitted, making it suitable for selection of nutritional markers as well as the study of biomass utilization by C. bescii. A low osmolarity complex growth medium (LOC) was developed that dramatically improves growth and culture viability during storage, making it a better medium for routine growth and passaging of C. bescii. Both media contain significantly lower solute concentration than previously published media, allowing for flexibility in developing more specialized media types while avoiding the issues of growth inhibition and cell lysis due to osmotic stress. Plating on LOD medium solidified by agar results in ~1,000-fold greater plating efficiency than previously reported and allows the isolation of discrete colonies. These new media represent a significant advance for both genetic manipulation and the study of biomass utilization in C. bescii, and may be applied broadly across the Caldicellulosiruptor genus.