PubMed · 13126397
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J A SIMPSON. 1954. Dermatological changes in hypocalcaemia.. https://doi.org/10.1111/j.1365-2133.1954.tb12552.x
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A novel high-capacity, high-efficiency DNA extraction method is described using a photopolymerized silica-based monolithic column in a fused-silica capillary. Development involved investigation of the composition of the sol-gel monomer, fabrication conditions, and surface modifications in order to optimize the binding capacity. Extraction capacity and efficiency with the 3-(trimethoxysilyl)propyl methacrylate (TMSPM) monolith formulations fabricated in capillaries were investigated using a simple three-step procedure consisting of sample loading, washing of the solid phase, and elution of the DNA using a low ionic strength Tris buffer at pH 8. Once the TMSPM monomer concentration was optimized to yield a monolith with maximum test stability (robustness) and minimum back pressure, the monolith surface was modified by the grafting of tetramethyl orthosilicate (TMOS) for increased DNA binding capacity. After the examination of a variety of TMOS concentrations, 85% v/v TMOS was found to be optimal for DNA extraction without any obvious changes to the monolith structure. The reduction of time allowed for TMSPM hydrolysis prior to UV polymerization from 20 to 5 min led to a lower back pressure of the monolith, enabling better TMOS derivatization and therefore higher binding capacity. Minimal buffer volume (as low as 1 muL) was required to elute DNA from the solid phase, providing a DNA concentrating effect potentially important for downstream processes. While experimentation employed monolithic columns that were 12 cm in length, reduction of the length to 2 cm still allowed for a DNA binding capacity of at least 100 ng of prepurified human genomic DNA and extraction efficiencies greater than 85%. Extraction of low sample volumes (submicroliter) of human whole blood were successfully performed, with extraction efficiencies from the 2-cm monolithic column higher than those obtained from a commercial DNA extraction kit. These results position this novel matrix as an attractive alternative for solid-phase extraction of DNA and other biologically active molecules in microscale devices.
Radiation safety was determined to maintain quality control in the cyclotron laboratory. Based on the results of 438 runs in the Faraday cup (20 microA for 10 min), 20 runs on 18O-water target (40 microA for 2 h) and 10 runs on 18O-gas targets (30 microA for 45 min), we have established that occupationally exposed workers remain 10 +/- 5 times below federal regulatory limits (FRLs) in the cyclotron vault, 30 +/- 8 times below FRL in the radiochemistry laboratory and 200 +/- 10 times below the FRL outside the cyclotron laboratory during beam operation. (The FRL for unrestricted area are <20 microSv in 1 h.) The non-occupationally exposed workers serving in offices in the vicinity of the cyclotron vault within 100 m distance remained 200 times below the FRL irrespective of beam being on or off, suggesting that routine beam operation of 40 microA for 2 h once a day during office hours is safe provided quality control and system performance measures as discussed in this report are strictly maintained.
Recovery of Leptospira in the clinical setting is typically low as specialized culture media is needed. Previous data demonstrated that blood culture media commonly available to most clinical laboratories do not adequately sustain viable Leptospira. We hypothesized that mycobacterial blood culture medium, which is often readily available to most clinical laboratories, might be able to support the growth of Leptospira. Leptospires and fresh human blood were inoculated into BacT/ALERT (bioMérieux, Durham NC) mycobacterial (MB) and enriched mycobacterial bottles. Standard aerobic (FA) and anaerobic (SN) bottles were also inoculated as a control group. Inoculated bottles were then evaluated for their ability to support Leptospira growth using dark-field microscopy, subculture, and an automated growth detection system. Viable leptospires were detected in MB bottles up to day 14. FA and SN were performed in accordance with prior data. We conclude that MB and enriched MB bottles of the BacT/ALERT blood culture system can support viable leptospires.