Efficient recovery of low endotoxin medium-chain-length poly([R]-3-hydroxyalkanoate) from bacterial biomass

Bacterial polyhydroxyalkanoate (PHA) is an attractive biopolyester for medical applications due to its biocompatibility. However, inappropriate extraction of PHA from bacterial biomass results in contamination by pyrogenic compounds (e.g. lipopolysaccharides) and thus influences medical testing. This problem was solved by a temperature-controlled method for the recovery of poly(3-hydroxyoctanoate-co-3-hydroxyhexanaote) (PHO) from Pseudomonas putida GPo1. In contrast to other methods, precipitation of PHO was triggered by cooling the hot solution to a particular temperature. N-hexane and 2-propanol were found to be optimal solvents for such procedure. Quantitative extraction with n-hexane took place at 50 degrees C and optimal precipitation occurred between 0 and 5 degrees C. The purity was >97% (w/w) and the endotoxicity between 10 and 15 EU/g PHO. Additional re-dissolution in 2-propanol at 45 degrees C and precipitation at 10 degrees C resulted in a purity of close to 100% (w/w) and the minimal endotoxicity of 2 EU/g PHO. The polydispersity (M(w)/M(n)) of PHO was decreased from 2.0 to 1.5 for this optimized procedure.     

The cellular prion protein and its role in Alzheimer disease

The cellular prion protein (PrP^C) is a membrane-bound glycoprotein especially abundant in the central nervous system (CNS). The scrapie prion protein (PrP^Sc, also termed prions) is responsible of transmissible spongiform encephalopathies (TSE), a group of neurodegenerative diseases which affect humans and other mammal species, although the presence of PrP^C is needed for the establishment and further evolution of prions.The present work compares the expression and localization of PrP^C between healthy human brains and those suffering from Alzheimer disease (AD).In both situations we have observed a rostrocaudal decrease in the amount of PrP^C within the CNS, both by immunoblotting and immunohistochemistry techniques. PrP^C is higher expressed in our control brains than in AD cases. There was a neuronal loss and astogliosis in our AD cases. There was a tendency of a lesser expression of PrP^C in AD cases than in healthy ones. And in AD cases, the intensity of the expression of the unglycosylated band is higher than the di- and monoglycosylated bands.With regards to amyloid plaques, those present in AD cases were positively labeled for PrP^C, a result which is further supported by the presence of PrP^C in the amyloid plaques of a transgenic line of mice mimicking AD.The work was done according to Helsinki Declaration of 1975, and approved by the Ethics Committee of the Faculty of Medicine of the University of Navarre.Key words: cellular prion protein, Alzheimer disease, transgenic mice     

How to monitor elusive lizards: comparison of capture–recapture methods on giant day geckos (Gekkonidae, Phelsuma madagascariensis grandis) in the Masoala rainforest exhibit, Zurich Zoo

Rapid and reliable estimation of population size is needed for the efficient monitoring of animal populations of conservation concern. Unfortunately, technical advances in this area have not been paralleled in uptake in conservation, which may be due to difficulties in implementation or the lack of general guidelines for application. Here we tested five different methods used to estimate population size [capture–mark–recapture (CMR), finite-mixture models, model averaging of finite-mixture models, accumulation curve methods (ACM), and the line transect method (LT)] using extensive capture–recapture data of the giant day gecko (Gekkonidae, Phelsuma madagascariensis grandis, Gray 1870) at the Masoala rainforest exhibit, Zurich Zoo. When the complete data were analyzed [30 sessions (and 27 sessions for the LT)], all methods except the LT produced similar estimates of population size. The simple ACM gave a small coefficient of variation (CV), but did not cover the most likely value of population size at moderate sampling effort. Nevertheless, the ACM was the only method that showed a reasonable convergence when subsets of data were used. CMR and Pledger models included the reference value in their confidence intervals (CI) after 25 and 30 sessions, respectively. Although model averaging did slightly improve the estimate, the CV was still high for the full dataset. Our method of using subsets of data to test the robustness of estimates is simple to apply and could be adopted more widely in such analyzes to evaluate sensitivity to method of evaluation. In conclusion, simple accumulation methods showed similar efficiency to more complex statistical models, and are likely to be sufficiently precise for most conservation monitoring purposes. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Schweizerische Vegetationsforschung von 1938 bis 1948

Ohne ZusammenfassungManuskript eingegangen am 27.X.1949.     

Optimizing Human Synovial Fluid Preparation for Two-Dimensional Gel Electrophoresis

Background

Prenatal screening for Down Syndrome (DS) would benefit from an increased number of biomarkers to improve sensitivity and specificity. Improving sensitivity and specificity would decrease the need for potentially risky invasive diagnostic procedures.

Results

We have performed an in depth two-dimensional difference gel electrophoresis (2D DIGE) study to identify potential biomarkers. We have used maternal plasma samples obtained from first and second trimesters from mothers carrying DS affected fetuses compared with mothers carrying normal fetuses. Plasma samples were albumin/IgG depleted and expanded pH ranges of pH 4.5 - 5.5, pH 5.3 - 6.5 and pH 6 - 9 were used for two-dimensional gel electrophoresis (2DE). We found no differentially expressed proteins in the first trimester between the two groups. Significant up-regulation of ceruloplasmin, inter-alpha-trypsin inhibitor heavy chain H4, complement proteins C1s subcomponent, C4-A, C5, and C9 and kininogen 1 were detected in the second trimester in maternal plasma samples where a DS affected fetus was being carried. However, ceruloplasmin could not be confirmed as being consistently up-regulated in DS affected pregnancies by Western blotting.

Conclusions

Despite the in depth 2DE approach used in this study the results underline the deficiencies of gel-based proteomics for detection of plasma biomarkers. Gel-free approaches may be more productive to increase the number of plasma biomarkers for DS for non-invasive prenatal screening and diagnosis.