Nonradioactive Northern blotting with biotinylated and digoxigenin-labeled RNA probes

The application of nonradioactive RNA probes for Northern blotting offers the advantage of a rapid turn-around time for results without the loss of sensitivity for target mRNA detection. However, a problem that has impeded the widespread use of nonradioactive RNA probes for use in Northern blotting is the difficulty in stripping these probes from nylon membranes after hybridization. In this report we describe two protocols for stripping digoxigenin (Dig)-labeled RNA probes from nylon membranes. One protocol utilizes a phosphate-buffered formamide stripping solution to remove nonchemically modified (regular) RNA probes while the other method utilizes strippable probes that were produced with a chemically modified nucleotide (CTP) and removed by a specific stripping solution. This latter method was developed by Ambion Inc. and is called Strip-EZ. We also describe a protocol for the detection of two separate rat mRNAs using both biotin and digoxigenin-labeled RNA probes that does not require stripping the membrane after hybridization. Finally, we describe the use of another new labeling technology, called Chem-Link, that quickly and conveniently labels RNA for use in Northern blotting.     

Viscoelastic behavior of highly crosslinked poly(acrylic acid)

The effects of crosslinking upon the dynamic mechanical properties and swelling behavior of poly(acrylic acid) (PAA) have been studied. Materials were prepared by the free radical copolymerization of acrylic acid with varying amounts of the tetrafunctional monomer allyl acrylate (ALA). The results indicated a linear dependence of the glass-transition temperature (T_g) on composition, T_g increasing by ～43°C over the mole fraction range X = 0?0.37 ALA. Room temperature (25°C) modulus values, as determined by both dynamic and compression methods, were inversely proportional to the initial concentration of ALA. The degree of network formation has been characterized in terms of the molecular weight between crosslinks M_c, and the influence of this parameter on the swelling ratio was discussed.     

Empirische Methodologien und Methoden der angewandten und der empirischen Ethik

Definition of the problem This paper deals with the question of adequate methodologies and methods to conduct empirical research and integrate empirical knowledge in the field of biomedical ethics. Arguments It starts with reflections about the main subject matter of ethics, the human being as a bio-psycho-social creature and its morals, and argues that social sciences are the adequate basic empirical science in the field of ethics in general. The next paragraph considers theory of science perspectives of the relationship of ethics, empirical findings, methodologies and theories and examples of the integration of empirical findings into ethical deliberations. Reasons for the tension between social science and ethics are scrutinized which are different in the yet to be defined field of empirical ethics compared to traditional applied ethics. Conclusion How a bio-psycho-social ethics that incorporates these criteria into its own body of work could look like is finally depicted.     

SEMPA – Ein Ansatz des Semantischen Prozessmanagements zur Planung von Prozessmodellen

Currently process modeling is mostly done manually. Therefore, the initial design of process models as well as changes to process models which are frequently necessary to react to new market developments or new regulations are time-consuming tasks. In this paper we introduce SEMPA, an approach for the partly automatic planning of process models. Using ontologies to semantically describe actions – as envisioned in Semantic Business Process Management –, a process model for a specified problem setting can be created automatically. In comparison to existing planning algorithms our approach creates process models including control structures and is able to cope with complex and numerical input and output parameters of actions. The prototypical implementation as well as an example taken from the financial services domain illustrate the practical benefit of our approach.     

Using physical memorabilia as opportunities to move into collocated digital photo-sharing

The uptake of digital photos vs. print photos has altered the practice of photo-sharing. Print photos are easy to share within the home, but much harder to share outside of it. The opposite is true of digital photos. People easily share digital photos outside the home, e.g., to family and friends by e-mail gift-giving, and to social networks and the broader public by web publishing. Yet within the home, collocated digital photo-sharing is harder, primarily because digital photos are typically stored on personal accounts in desktop computers located in home offices. This leads to several consequences. (1) The invisibility of digital photos implies few opportunities for serendipitous photo-sharing. (2) Access control and navigation issues inhibit family members from retrieving photo collections. (3) Photo viewing is compromised as digital photos are displayed on small screens in an uncomfortable viewing setting.To mitigate some of these difficulties, we explore how physical memorabilia collected by family members can create opportunities that encourage social and collocated digital photo-sharing. First, we studied (via contextual interviews with 20 households) how families currently practice photo-sharing and how they keep memorabilia. We identified classes of memorabilia that can serve as memory triggers to family events, trips, and times when people took photos. Second, we designed Souvenirs, a photo-viewing system that exploits memorabilia as a social instrument. Using Souvenirs, a family member can meaningfully associate physical memorabilia with particular photo-sets. Later, any family member can begin their story-telling with others through the physical memento, and then enrich the story by displaying its associated photos simply by moving the memento close to the home's large-format television screen. Third, we re-examined our design premises by evoking household reactions to an early version of Souvenirs. Based on these interviews, we redesigned Souvenirs to better reflect the preferences and real practices of photo and memorabilia use in the home.     

Viral interleukin 10 (IL-10), the human herpes virus 4 cellular IL-10 homologue, induces local anergy to allogeneic and syngeneic tumors

After the cloning of murine cytokine synthesis inhibitory factor, it was recognized that a homologous open reading frame was encoded within the Epstein-Barr virus (human herpes virus 4). This viral protein has now been termed viral interleukin 10 (vIL-10) to reflect its protein sequence homology to "cellular" IL-10 (cIL-10, either murine or human IL-10). It is now widely accepted that vIL-10 shares many functions with cIL-10, principally, the ability to enhance survival of newly infected B cells and to diminish the production of IFN-gamma and IL-2 during ongoing immune reactions. The immunomodulatory effect of locally secreted vIL-10 and murine IL-10 (mIL-10) was examined in tumor models using CL8-1 (a BL6 melanoma cell line transfected with the H-2Kb class I gene) in syngeneic animals. Although parental BL6 tumor cells grow in immunocompetent syngeneic hosts, CL8-1 are rejected. To achieve local secretion of vIL-10, we generated vIL-10 retroviral vectors. While nontransduced CL8-1 cells (1 x 10(4)) failed to grow when injected intradermally in C57BL/6 mice, CL8-1 cells (1 x 10(4)) transduced with vIL-10 formed palpable tumors and eventually killed 80% of injected animals. Suppression of tumor rejection was also noted when CL8-1 tumors with or without vIL-10 transfection were admixed with syngeneic vIL-10-transfected fibroblasts and inoculated. Since the in vitro proliferation of the tumor was not altered after transduction with the vIL-10 gene and injection of vIL-10-transduced CL8-1 does not affect the rejection of nontransduced CL8-1 inoculated at a distant site, local vIL-10 secretion appears to suppress the process of immune rejection of the target cells in a dose-dependent manner. Similar results were observed for the H-2b MCA105 sarcoma tumor model in allogeneic BALB/c mice (H-2d). Although all animals that received nontransfected MCA105 rapidly rejected these tumors, MCA105 sarcomas transfected with vIL-10 remained palpable for up to 37 d. The local immunosuppressive effect of gene-delivered vIL-10 could be neutralized by anti-human IL-10 monoclonal antibody or could be reversed by the systemic administration of IL-2 or IL-12. In marked contrast, mIL-10 transfection of CL8-1 significantly suppressed tumor growth and frequently led to the rejection of tumor. Similar results were obtained for the murine tumor cell lines MCA102.(ABSTRACT TRUNCATED AT 400 WORDS)     

Conformations and folding of lysozyme ions in vacuo

Proton transfer reactivity of isolated charge states of the protein hen egg-white lysozyme shows that multiple distinct conformations of this protein are stable in the gas phase. The reactivities of the 9+ and 10+ charge state ions, formed by electrospray ionization of "native" (disulfide-intact) and "denatured" (disulfide-reduced) solutions, are consistent with values calculated for ions in their crystal structure and fully denatured conformations, respectively. Charge states below 8+ of both forms, formed by proton stripping, have similar or indistinguishable reactivities, indicating that the disulfide-reduced ions fold in the gas phase to a more compact conformation.     

Obturation of a C-shaped canal using an improved method of warm lateral condensation

Pilot studies conducted by the authors seem to indicate that obturation of a root canal system by the lateral condensation of gutta-percha using an electrically heated spreader results in a more homogeneous mass of gutta-percha than standard lateral condensation. This article describes an improved variation which prevents inadvertent dislodgement of the mass of gutta-percha during the obturation. The authors refer to this technique as "zap and tap" and a case report describes its use to obturate a C-shaped canal system of a mandibular second molar.