Evaluation of real-time PCR results at the limit of detection

Real-time polymerase chain reaction (real-time PCR) facilitates to detect DNA fragments at very low copy numbers. Positive results, e.g. for unauthorized genetically modified organism (GMO) contamination or for allergens, may raise safety concerns and have far-reaching consequences. However, in case of very low concentrations of DNA samples, results for the same product lot or even for identical samples from different laboratories may differ. Therefore, an approach for a standardized interpretation and reporting of results obtained by real-time PCR at the limit of detection (LOD) is proposed. A quality control DNA sample containing the target at the LOD (95?%) is analysed in parallel with the real DNA sample and the respective C _T values are compared. In addition, practical approaches for in house and precision-based estimation of the LOD are presented. The proposed approach may also contribute to the current discussion on implementing a technical solution to handle DNA traces in specimen, e.g. for the detection of unauthorized GMO.     

Dynamische Prozeßsimulation in der industriellen Praxis

Dynamic process simulation in industrial practice. Dynamic process simulation is gaining in importance in process development and plant design. In addition to simulation tools for steady state calculations and optimization, recent years have seen increased development of software tools suitable for dynamic simulation. In particular, dynamic process models have been developed for various unit operations at universities and in industry. Application of the models includes synthesis and analysis of the control structure, start-up and shut down, safety assessments, optimization, and training. In addition, dynamic flowsheet programs analogous to the steady state programs were developed. Wider use of these program systems requires increased flexibility, solving procedures, user interfaces, model library, interfaces to external programs, etc. The present article is intended not only to describe the state of the art and describe some typical applications, but in particular to examine the demands placed on dynamic process simulation and the simulators and to delineate the current limitations of dynamic process simulation from an industrial viewpoint.     

弱渗透松散和坚硬岩石的渗透性测定方法

固体有害废物安全填埋场基础弱渗透岩石的渗透性,大多数情况下都由实验室和野外方法测定．测定方法的适用性取决于岩石种类、岩体状态和渗透性数量级别．在同一实验段内采用不同的实验方法,一情况下导致产生两个数量级的误差,而更大的不可靠性与使用不同的评价方法有关．因为可靠并可引用的渗透性测定值在废物安全填埋场建设中对地质屏障的评价具有重大意义,如果各地能同时列出科研规划,共同研究对所有可能存在的地质屏障岩石进行全面分析评价,确定不同实验方法之间相关关系,以及使评价方法尽可能统一,是十分必要的     

Synthesis and isolation of <Emphasis Type="Italic">trans</Emphasis>-7, <Emphasis Type="Italic">cis</Emphasis>-9 octadecadienoic acid and other CLA isomers by base conjugation of partially hydrogenated γ-linolenic acid

CLA is of considerable interest because of reported potentially beneficial effects in animal studies. CLA, while not yet unambiguously defined, is a mixture of octadecadienoic acids with conjugated double bonds. The major isomer in natural products is generally considered to be cis-9,trans-11-octadecadienoic acid (c9, t11), which represents >75% of the total CLA in most cases. Other isomers are drawing increased attention. The t7,c9 isomer, which is often the second-most prevalent CLA in natural products, has been reported to represent as much as 40% of total CLA in milk from cows fed a high-fat diet. The need for a reference material became apparent in a recent study directed specifically at measuring t7,c9-CLA in milk, plasma, and rumen. A suitable standard mixture was produced by stirring 0.5 g of γ-linolenic acid (all cis-6,9, 12-C_18∶3) with 100 mL of 10% hydrazine hydrate in methanol for 2.5 h at 45°C. The solution was diluted with H₂O and acidified with HCl. The resulting partially hydrogenated FA were extracted with ether/petroleum ether, dried with Na₂SO₄, and conjugated by adding of 6.6% KOH in ethlylene glycol and heating for 1.5 h at 150–160°C. Approximately 20 mg each of cis-6, trans-8; trans-7, cis-9; cis-9, trans-11; and rans-10, cis-12 were obtained along with other FA. Methyl esters (FAME) of these four cis/trans isomers were resolved by Ag⁺HPLC (UV 233) and partially resolved by GC/(MS or FID) (CP-Sil 88). Treatment of these FAME with I₂ yielded all possible cis/trans (geometric) isomers for the four positions 6,8; 7,9; 9,11; and 10,12.     

Application of the Electrodiffusion Method to Measure Wall Shear Stress: Integrating Theory and Practice

The electrodiffusion method has been used in fluid dynamic research for the past 50 years. It allows the measurement of wall shear stress, a crucial parameter, e.g., for the cleaning of membrane modules used in water filtration. Various authors have published articles dealing with the theory behind this technique. But no paper collects all the knowledge assembled over five decades of application. Here, comprehensive summary of the theory of steady flow, unsteady flow, and transient voltage step experiments is given. Factors influencing the accuracy of the measurements are discussed. Furthermore, a new approach to calibrate the system from voltage step experiments is introduced, and practical issues related to its application in flow measurements are discussed for an exemplary signal response to a near‐wall flow.     

Mitotic Diversity in Homeostatic Human Interfollicular Epidermis

Despite decades of skin research, regulation of proliferation and homeostasis in human epidermis is still insufficiently understood. To address the role of mitoses in tissue regulation, we utilized human long-term skin equivalents and systematically assessed mitoses during early epidermal development and long-term epidermal regeneration. We now demonstrate four different orientations: (1) horizontal, i.e., parallel to the basement membrane (BM) and suggestive of symmetric divisions; (2) oblique with an angle of 45°–70°; or (3) perpendicular, suggestive of asymmetric division. In addition, we demonstrate a fourth substantial fraction of suprabasal mitoses, many of which are committed to differentiation (Keratin K10-positive). As verified also for normal human skin, this spatial mitotic organization is part of the regulatory program of human epidermal tissue homeostasis. As a potential marker for asymmetric division, we investigated for Numb and found that it was evenly spread in almost all undifferentiated keratinocytes, but indeed asymmetrically distributed in some mitoses and particularly frequent under differentiation-repressing low-calcium conditions. Numb deletion (stable knockdown by CRISPR/Cas9), however, did not affect proliferation, neither in a three-day follow up study by life cell imaging nor during a 14-day culture period, suggesting that Numb is not essential for the general control of keratinocyte division.     

Nanoparticle evolution in flame spray pyrolysis—Process design via experimental and computational analysis

In flame spray pyrolysis (FSP), the evolution of metal oxide nanoparticles relies on quite a number of droplet (liquid) and vapor phase related physical mechanism as for instance precursor evaporation, oxidation, nucleation via gas-to-particle conversion mechanism, and subsequent particle (solid) growth mechanisms based on coagulation, sintering/coalescence, and agglomeration. The liquid precursor and dispersion oxygen feed rates are relevant control parameters of the FSP process for tailoring the nanoparticle size (diameter) and structure as well as the atomizer nozzle configuration. Sophisticated nonintrusive, laser-based in situ and ex situ diagnostics with multiscale spatial resolution (micrometer to meter range) are applied for analyzing droplet formation and size, gas velocity, temperature, species concentration, as well as primary and agglomerate diameters along the flow direction. Computational fluid dynamics (CFD) are coupled with population balance modeling (PBM) to elucidate the nanoparticle dynamics within the reactive spray. It is found that the CFD-PBM approach allows estimations of primary and agglomerate nanoparticle diameters within 80 and 75% accuracy compared to experimental data, suggesting that the methods presented could pave the way for designing next-generations of flame reactors.     

Carborane-Containing Matrix Metalloprotease (MMP) Ligands as Candidates for Boron Neutron-Capture Therapy (BNCT)

Based on the previously reported potent and selective sulfone hydroxamate inhibitors SC-76276, SC-78080 (SD-2590), and SC-77964, potent MMP inhibitors have been designed and synthesized to append a boron-rich carborane cluster by employing click chemistry to target tumor cells that are known to upregulate gelatinases. Docking against MMP-2 suggests binding involving the hydroxamate zinc-binding group, key H-bonds by the sulfone moiety with the peptide backbone residues Leu82 and Leu83, and a hydrophobic interaction with the deep P1’ pocket. The more potent of the two triazole regioisomers exhibits an IC₅₀ of 3.7 nM versus MMP-2 and IC₅₀ of 46 nM versus MMP-9.