An improved silver staining procedure for schizodeme analysis in polyacrylamide gradient gels

A simple protocol is described for the silver staining of polyacrylamide gradient gels used for the separation of restriction fragments of kinetoplast DNA [schizodeme analysis of trypanosomatids (Morel et al., 1980)]. The method overcomes the problems of non-uniform staining and strong background color which are frequently encountered when conventional protocols for silver staining of linear gels. The method described has proven to be of general applicability for DNA, RNA and protein separations in gradient gels.

Modelling dependence in a ratemaking procedure with multivariate Poisson regression models

When actuaries face with the problem of pricing an insurance contract that contains different types of coverage, such as a motor insurance or homeowner's insurance policy, they usually assume that types of claim are independent. However, this assumption may not be realistic: several studies have shown that there is a positive correlation between types of claim. Here we introduce different regression models in order to relax the independence assumption, including zero-inflated models to account for excess of zeros and overdispersion. These models have been largely ignored to multivariate Poisson date, mainly because of their computational di±culties. Bayesian inference based on MCMC helps to solve this problem (and also lets us derive, for several quantities of interest, posterior summaries to account for uncertainty). Finally, these models are applied to an automobile insurance claims database with three different types of claims. We analyse the consequences for pure and loaded premiums when the independence assumption is relaxed by using different multivariate Poisson regression models and their zero-inflated versions.

A mixed splicing procedure for economic time series

This note develops a flexible methodology for splicing economic time series that avoids the extreme assumptions implicit in the procedures most commonly used in the literature. It allows the user to split the required correction to the older of the series being linked between its levels and growth rates on the basis what he knows or conjectures about the persistence of the factors that account for the discrepancy between the two series that emerges at their linking point. The time profile of the correction is derived from the assumption that the error in the older series reflects the inadequate coverage of emerging sectors or activities that grow faster than the aggregate.

A homing procedure for studying spatial memory in immature and adult rodents

In this procedure, subjects learn the spatial position of one hole out of many, that allows them to escape from a large open-field into their home cage. The arena is circular and can be rotated between trials so that no proximal landmark is permanently associated with the target hole. This task is thus similar to the Morris water maze procedure, since subjects must remember the position of the escape hole relative to extra-arena cues only. In addition it allows studying the importance of olfactory cues such as scent marks in or around a hole. Since the motivation is to reach home and the motor requirement is low, this task provides a useful alternative to the Morris place navigation task for studying spatial orientation in weanling or senescent rats. Examples are given showing that various behavioural parameters provide a good estimation as how subjects learn this task.

Adaptable pulmonary artery band for late arterial switch procedure in transposition of the great arteries.

BACKGROUND: In late-diagnosed transposition of the great arteries (TGA), the left ventricle (LV) involutes as it pumps against low resistance and needs retraining by applying a pulmonary artery band (PAB) in preparation for an arterial switch operation. We report our experience with a telemetrically adaptable band compared with classic banding. METHODS: Ten patients underwent retraining of the LV, 4 patients with an adaptable band and progressive weekly tightening of the band (group 1) and 6 patients with a traditional band (group 2). RESULTS: Mean weight and age at pulmonary band placement was 5.8 ± 2.36 kg and 11.7 ± 11.1 months for group 1 and 5.0 ± 2.3 kg and 6.4 ± 7.6 months for group 2. Time between palliation and switch procedure was 4.2 months in both groups. Group 1 showed an initial mean pulmonary gradient of 25.5 ± 4.43 mm Hg with a 5% closure of the device. The mean gradient increased with progressive closure to 63.5 ± 9.8 mm Hg at the time of the arterial switch operation. There were no reinterventions or deaths in this group. In group 2, the mean pulmonary gradient increased with growth from 49 ± 21.4 mm Hg to 68.4 ± 7.86 mm Hg at the time of the switch procedure. However, 4 of these patients required reoperations during retraining: 2 needed 1 reoperation and 2 needed 2 reoperations. Two patients died-1 after banding and 1 after the switch operation. CONCLUSIONS: Retraining of the LV by the adaptable device allows precise control of the tightening, avoids repetitive operations, and diminishes morbidity.

111In-pentetreotide scintigraphy: procedure guidelines for tumour imaging.

This document provides general information about somatostatin receptor scintigraphy with (111)In-pentetreotide. This guideline should not be regarded as the only approach to visualise tumours expressing somatostatin receptors or as exclusive of other nuclear medicine procedures useful to obtain comparable results. The aim of this guideline is to assist nuclear medicine physicians in recommending, performing, reporting and interpreting the results of (111)In-pentetreotide scintigraphy.

TT virus infection in children and adults who visited a general hospital in the south of Brazil for routine procedure

TT virus (TTV) is a newly described nonenveloped human virus, with a circular, negative-stranded DNA genome, that was first identified in the blood of a patient with posttransfusion hepatitis of unknown etiology. PCR primers and conditions used for TTV DNA amplification may greatly influence the level of TTV detection in serum. Three PCR assays, with different regions of the genome as targets, were used to test TTV DNA in 130 sera from children and adults visiting a hospital in the south of Brazil, most of them for routine procedure. Forty-four percent of adult sera and 73% of sera from children aged 0-10 years were TTV positive with at least one PCR assay. However, the three assays were able to detect only 33%, 35%, and 70% of the total positive samples. Our results showed a high prevalence of TTV infection in the south of Brazil, particularly among young children, and confirmed the necessity of performing several PCR assays to assess the true TTV prevalence in a determined population.

Inhibition of the polymerase chain reaction by sputum samples from tuberculosis patients after processing using a silica-guanidiniumthiocyanate DNA isolation procedure

With the objective to evaluate PCR-mediated detection of Mycobacterium tuberculosis DNA as a diagnostic procedure for diagnosis of tuberculosis in individuals attending ambulatory services in Primary Health Units of the City Tuberculosis Program in Rio de Janeiro, Brazil, their sputum samples were collected and treated with a DNA extraction procedure using silica-guanidiniumthiocyanate. This procedure has been described to be highly efficient for extraction of different kind of nucleic acids from bacteria and clinical samples. Upon comparing PCR results with the number of acid-fast bacilli, no direct relation was observed between the number of bacilli present in the sample and PCR positivity. Part of the processed samples was therefore spiked with pure DNA of M. tuberculosis and inhibition of the PCR reaction was verified in 22 out of 36 (61%) of the samples, demonstrating that the extraction procedure as originally described should not be used for PCR analysis of sputum samples.

Multiple nodules of the scrotum: histopathological findings and surgical procedure. A study of five cases.

BACKGROUND: Multiple nodules of the scrotum are uncommonly reported. Their origin is controversial. Treatment is always surgical but the best procedure is still to be determined. MATERIALS AND METHODS: Five new cases are reported with description of the histopathological findings and surgical procedure. RESULTS: Nodules of the scrotum were more frequent in patients with dark skin suggesting an ethnic susceptibility. No other predisposing factors were noted. Screening for disturbances of phosphate or calcium balance was negative. The following histopathological findings were observed: non-calcified epidermoid cysts (3 patients), calcified epidermoid cysts (1 patient) and nodular calcifications without epithelial or glandular structures (1 patient). Subtotal excisions of the scrotum wall using tumescent anaesthesia were performed in all patients without any significant complications. Cosmetic results were excellent. No new lesions were observed during the 1-year follow-up period. CONCLUSIONS: Most cases of multiple nodules of the scrotum are due to non-calcified epidermoid cysts. The term scrotal calcinosis is therefore probably abusively used by many authors. Some cases of nodular calcifications may be due to dystrophic calcification of epidermoid cysts, but calcifications may also occur without any visible epithelial or glandular structure. Subtotal excision of the scrotum wall is a safe and effective surgical procedure to treat multiple nodules of the scrotum. Cosmetic results are excellent and recurrences are rare.

Quantification of nicotine, cotinine, trans-3'-hydroxycotinine and varenicline in human plasma by a sensitive and specific UPLC-tandem mass-spectrometry procedure for a clinical study on smoking cessation.

A sensitive and specific ultra performance liquid chromatography-tandem mass spectrometry method for the simultaneous quantification of nicotine, its metabolites cotinine and trans-3'-hydroxycotinine and varenicline in human plasma was developed and validated. Sample preparation was realized by solid phase extraction of the target compounds and of the internal standards (nicotine-d4, cotinine-d3, trans-3'-hydroxycotinine-d3 and CP-533,633, a structural analog of varenicline) from 0.5mL of plasma, using a mixed-mode cation exchange support. Chromatographic separations were performed on a hydrophilic interaction liquid chromatography column (HILIC BEH 2.1×100mm, 1.7μm). A gradient program was used, with a 10mM ammonium formate buffer pH 3/acetonitrile mobile phase at a flow of 0.4mL/min. The compounds were detected on a triple quadrupole mass spectrometer, operated with an electrospray interface in positive ionization mode and quantification was performed using multiple reaction monitoring. Matrix effects were quantitatively evaluated with success, with coefficients of variation inferior to 8%. The procedure was fully validated according to Food and Drug Administration guidelines and to Société Française des Sciences et Techniques Pharmaceutiques. The concentration range was 2-500ng/mL for nicotine, 1-1000ng/mL for cotinine, 2-1000ng/mL for trans-3'-hydroxycotinine and 1-500ng/mL for varenicline, according to levels usually measured in plasma. Trueness (86.2-113.6%), repeatability (1.9-12.3%) and intermediate precision (4.4-15.9%) were found to be satisfactory, as well as stability in plasma. The procedure was successfully used to quantify nicotine, its metabolites and varenicline in more than 400 plasma samples from participants in a clinical study on smoking cessation.