Development of an antipeptide antibody that binds to the C-terminal region of human CYP1B1

An antipeptide antibody was raised against a 14-mer synthetic peptide (CDFRANPNEPA KMN) corresponding to the amino acid sequence from 491 to 504 of human cytochrome P-450 (CYP)1B1. Rabbit-derived antisera demonstrated the ability to induce moderately high antibody titers (>1:10(5)) as judged by enzyme-linked immunosorbent assay. In Western blot analysis, the purified antibody recognized a single protein band (estimated as 56 kDa) in microsomes prepared from human and rodent tissues. No significant cross-reactivity to either human CYP1A1 or human CYP1A2 protein was detected. Titration studies using recombinant human CYP1B1 and an enhanced chemiluminescence-based detection method demonstrated a minimal detection sensitivity for this antiserum at about 0.34 ng/band in 8 x 7-cm minigels. The immunoprecipitation and immunoinhibition results indicate that this antisera recognizes the nondenatured human CYP1B1 protein but does not inhibit its enzyme activity. Using this antibody, CYP1B1 protein was detected in nine different human tissues and in cultured cells induced by various chemicals. This highly specific, highly sensitive antibody provides an important tool to study tissue distribution and cellular expression levels of CYP1B1, with negligible cross-reactivity from the other members of the CYP1 family.     

Free leptin, bound leptin, and soluble leptin receptor in normal and diabetic pregnancies

We measured serum levels of free leptin, bound leptin, and soluble leptin receptor by specific RIA methods in 20 normal and 19 insulin-dependent diabetes mellitus subjects at 20 and 30 weeks gestation and postpartum, and analyzed the data using hierarchical statistical models. Total leptin levels rise from 20-30 weeks gestation (688 +/- 58 to 785 +/- 62 pmol/L, mean +/- SEM; P = 0.009). There is a significant postpartum fall to 445 +/- 47 pmol/L (P < 0.001). This rise is caused by the rise in the bound leptin levels, as there is no significant change in free leptin levels between 20 and 30 weeks (P = 0.17). There is a significant postpartum fall in free leptin levels (P < 0.001). Insulin requirements rise in the third trimester, but despite this there was no significant difference in free or bound leptin levels between the normal and diabetic subjects at any stage [free leptin, 223 +/- 35 and 266 +/- 24, 237 +/- 45 and 223 +/- 27, and 109 +/- 16 and 104 +/- 24 (P = 0.34); bound leptin, 410 +/- 73 and 428 +/- 54, 501 +/- 78 and 562 +/- 71, and 330 +/- 47 and 271 +/- 46 (P = 0.84); for normals and diabetics at 20 and 30 weeks gestation and postpartum, respectively]. Diabetic subjects, however, had significantly higher soluble leptin receptor levels at all stages (P < 0.001), which rose further in the third trimester from 3742 +/- 268 (mean +/- SEM) to 4134 +/- 239 pmol/L, whereas in the normal group there was a fall from 3149 +/- 169 to 2712 +/- 123 (P = 0.05). There is a linear relationship between the soluble leptin receptor levels and the body mass index in the diabetic group only. We conclude that there is no significant difference in free or bound leptin levels between the normal and insulin-dependent diabetic subjects either during pregnancy or postpartum, but female insulin-dependent diabetic subjects have significantly higher soluble leptin receptor levels. We speculate that high soluble leptin receptor levels might be implicated in the development of the leptin resistance in this group.     

Tasks of the NRLs with respect to crisis management: contribution of the EURL/NRL at the BVL as exemplified by the horse meat scandal

According to Art. 32 and 33 of Regulation (EC) No. 882/2004, the European Union and National Reference Laboratory for Pharmacologically Active Substances (EURL/NRL) at the Federal Office of Consumer Protection and Food Safety (BVL) is responsible, among other tasks, for supporting the official control laboratories as well as the competent authorities on a national and EU level. It is thus an important instrument in the overall complex of crisis management. Immediately after the publication of the first rapid alerts in the framework of the European Rapid Alert System for Food and Feed (RASFF), the EURL provided advice to the responsible Commission service with respect to the wording of the Commission Recommendation of 19 February 2013 on a coordinated control plan with a view to establish the prevalence of fraudulent practices in the marketing of certain foods, by detailing the available analytical means and capacities. In connection with the consumer fraud regarding the undeclared presence of horse meat in beef products (convenience food), a possible risk emanating from the non-authorised use of phenylbutazone (PBZ) was taken into consideration. For this reason the Commission Recommendation of 19 February 2013 suggested that at least one sample for every 50 tonnes of horse meat should be tested, each Member State (MS) being required to carry out a minimum of five tests. Moreover there was need for an analysis for PBZ in processed products containing horse meat as undeclared ingredient. The EURL/NRL of the BVL supported the analytical activities of the German control laboratories as well as of the European National Reference Laboratories (NRLs) through numerous measures.     

Development of Ni–Cu Materials Library by Using Combinatorial Pulsed Electrodeposition

Combinatorial materials fabrication and high-throughput characterization methods offer a new experimental paradigm to accelerate the enhancement of known and the discovery of new materials in materials science and engineering. In this study, the composition spread library of binary Ni–Cu alloy system is synthesized combinatorially onto a copper substrate using pulsed electrodeposition (PED) from a single sulfate bath with a complexing agent trisodium citrate in a modified Hull cell. Different current densities are expected on the tilted cathode which varied the composition along lateral direction and generated Ni–Cu binary spread. Crystallographic structure as well as atomic concentration of the constituent elements of the deposited binary Ni–Cu alloy film along the lateral direction is determined as a function of fabrication parameters. The presented results indicate the successful development of Ni–Cu binary spread (10–90 at.%) via PED.     

Histone composition of nucleosomes isolated from cultured Chinese hamster cells

Nuclei isolated from cultured Chinese hamster cells were treated with micrococcal nuclease and lysed, and the resulting chromatin subunit classes (nucleosomes) were purified by sedimentation and resedimentation through isokinetic sucrose gradients. Nucleosomes isolated from [3H]thymidine-labeled cells were analyzed for DNA size using both polyacrylamide gel and electron microscopic techniques. Nucleosomes isolated from [14C]lysine-labeled cells were analyzed for protein content using a sodium dodecyl sulfate-polyacrylamide gel system. The results from monitoring the [14c]lysine in each protein indicate that, in the nucleosome classes (monomer through tetramer), the molar ratios of histones H2A, H2B, H3, and H4 are equivalent. Furthermore, in each population of the nucleosome classes monomer through tetramer, it was possible to demonstrate that this histone unit (H2A + H2B + H3 + H4) is present, on the average, in the amount of two for monomers, four for dimers, six for trimers, and eight for tetramers. This is direct experimental confirmation of the prediction of R.D. Kornberg [(1974) Science 184, 868] concerning the substructure of chromatin.     

Book reviews

The chronology of various economies based on differing energy‐carriers is examined within a historic perspective and from an environmental point of view. The ‘Methanol Economy’ may well prove to be a permanent, yet ever‐increasing, under‐current of these other economies, each of which plays a temporary role on the Stage of Human History. Although current methods of methanol (MeOH) production are not as profitable as those of fossil‐ and bio‐fuels, scientific discovery, environmental concerns, dwindling resources and increased demand are certain to change their relative profitabilities. The reduced energy content of methanol, relative to gasoline or ethanol, is sure to be offset by the increase in technological efficiency of the machines being fuelled. Any potential hazards associated with the use of methanol can be tackled through education. In all, the production and use problems associated with methanol are outweighed by its benefits. These reduce dependence on fossil fuel.     

On-road bicycle facilities and bicycle crashes in Iowa, 2007–2010

Purpose

An average of 611 deaths and over 47,000 bicyclists are injured in traffic-related crashes in the United States each year. Efforts to increase bicycle safety are needed to reduce and prevent injuries and fatalities, especially as trends indicate that ridership is increasing rapidly. The objective of this study was to evaluate the effect of bicycle-specific roadway facilities (e.g., signage and bicycle lanes) in reducing bicycle crashes.

Methods

We conducted a case site-control site study of 147 bicycle crash-sites identified from the Iowa Department of Transportation crash database from 2007 to 2010 and 147 matched non-crash sites. Control sites were randomly selected from intersections matched to case sites on neighborhood (census block group) and road classification (arterial, feeder, collector, etc.). We examined crash risk by any on-road bicycle facility present and by facility type (pavement markings--bicycle lanes and shared lane arrows, bicycle-specific signage, and the combination of markings and signage), controlling for bicycle volume, motor vehicle volume, street width, sidewalks, and traffic controls.

Results

A total of 11.6% of case sites and 15.0% of controls had an on-road bicycle facility. Case intersections had higher bicycle volume (3.52 vs. 3.34 per 30 min) and motor vehicle volume (248.77 vs. 205.76 per 30 min) than controls. Our results are suggestive that the presence of an on-road bicycle facility decreases crash risk by as much as 60% with a bicycle lane or shared lane arrow (OR = 0.40, 95% CI = 0.09–1.82) and 38% with bicycle-specific signage (OR = 0.62, 95% CI = 0.15–2.58).

Conclusions

Investments in bicycle-specific pavement markings and signage have been shown to be beneficial to traffic flow, and our results suggest that they may also reduce the number of bicycle-motor vehicle crashes and subsequent injuries and fatalities. As a relatively low-cost traffic feature, community considerations for further implementation of these facilities are justified.