The Interlaboratory RObustness of Next-generation sequencing (IRON) study: a deep sequencing investigation of TET2, CBL and KRAS mutations by an international consortium involving 10 laboratories.

Massively parallel pyrosequencing allows sensitive deep sequencing to detect molecular aberrations. Thus far, data are limited on the technical performance in a clinical diagnostic setting. Here, we investigated as an international consortium the robustness, precision and reproducibility of amplicon next-generation deep sequencing across 10 laboratories in eight countries. In a cohort of 18 chronic myelomonocytic leukemia patients, mutational analyses were performed on TET2, a frequently mutated gene in myeloproliferative neoplasms. Additionally, hotspot regions of CBL and KRAS were investigated. The study was executed using GS FLX sequencing instruments and the small volume 454 Life Sciences Titanium emulsion PCR setup. We report a high concordance in mutation detection across all laboratories, including a robust detection of novel variants, which were undetected by standard Sanger sequencing. The sensitivity to detect low-level variants present with as low as 1-2% frequency, compared with the 20% threshold for Sanger-based sequencing is increased. Together with the output of high-quality long reads and fast run time, we demonstrate the utility of deep sequencing in clinical applications. In conclusion, this multicenter analysis demonstrated that amplicon-based deep sequencing is technically feasible, achieves high concordance across multiple laboratories and allows a broad and in-depth molecular characterization of cancer specimens with high diagnostic sensitivity.

[Changes in treatment of squamous cell carcinoma over time. A process analysis]. / Adaptierte Behandlungsmodalitäten bei Patienten mit Plattenepithelkarzinomen im Wandel der Zeit. Eine Prozessanalyse.

BACKGROUND: The incidence of squamous cell carcinoma is rapidly increasing and requires process-optimized treatment dependent on the course of the patient's individual disease. Patient-based data on squamous cell carcinoma were used to analyze the treatment modalities before DRGs were introduced, after their introduction and after discussion about process-optimization in context of clinical pathways. PATIENTS AND METHODS: From the recorded data in the department of dermatology, all squamous cell carcinoma cases were identified and evaluated. In addition to patient characteristics, parameters describing the process, for example, length of stay in hospital, pre- and post-interventional days, were collected. RESULTS: Between 1998-2002, 2004-2006 and January to June 2007, 658 patients were treated for squamous cell carcinoma. In contrast to the times before DRGs, the treatment process has been significantly optimized, reducing the pre- and post-operative days and thus the duration of stay. Analysis of clinical pathways as part of the continuous improvement process was hardly able to detect further improvement. However, the weekly distribution of the admitted patients has been improved. CONCLUSIONS: Patients with squamous cell carcinoma requiring surgical excision benefit from an optimized process, just as do impatient facilities. However, optimized treatment modalities carry hardly any potential of improvement by the invention of clinical pathways.

Continuous glucose monitoring by means of fiber-based, mid-infrared laser spectroscopy.

The application of mid-infrared laser spectroscopy to the reagent-free quantification of the concentration of glucose was investigated using cryogenically cooled lead salt lasers or, alternatively, quantum cascade lasers operating at room temperature. The concentration of glucose in aqueous solutions was quantified by means of fiber-based attenuated total reflection (ATR) spectroscopy (fiber-optical evanescent field analysis, FEFA) as well as fiber-based transmission spectroscopy. Both methods have the potential to be utilized by small fiber sensors, which can be inserted into the subcutaneous tissue in order to continuously measure the local concentration of glucose. In our in vitro experiments, noise-equivalent concentrations as low as 10 mg/dL were achieved. The mid-term stability of the measurement schemes was investigated by means of Allan variance analysis. Based on the research presented in this manuscript, an all-room-temperature measurement scheme using quantum cascade lasers, miniaturized fiber-optic sensors, and pyroelectric detectors appears well suited for the continuous monitoring of glucose concentrations at physiological levels.

DOAS measurements of tropospheric bromine oxide in mid-latitudes

Episodes of elevated bromine oxide (BrO) concentration are known to occur at high latitudes in the Arctic boundary layer and to lead to catalytic destruction of ozone at those latitudes; these events have not been observed at lower latitudes. With the use of differential optical absorption spectroscopy (DOAS), locally high BrO concentrations were observed at mid-latitudes at the Dead Sea, Israel, during spring 1997. Mixing ratios peaked daily at around 80 parts per trillion around noon and were correlated with low boundary-layer ozone mixing ratios.