Prokaryotic expression, purification and immunogenicity in rabbits of the small antigen of hepatitis delta virus

Funding Information: Expression and purification of HDV antigen was supported by Russian Foundation for Basic Research (grant 16-04-01490a). Evaluation of serum by Western blot and confocal microscopy was supported by Russian Science Foundation (grant 14-14-01021). Experiments in rabbits were supported by the Swedish Institute grants 09272_2013 and 19806_2016. Cross-border collaboration of the partners, exchange of the materials and standard operation procedures used in the study, and dissemination of the data were supported by the EU Twinning project VACTRAIN, contract nr 692293. Publisher Copyright: © 2016 by the authors; licensee MDPI, Basel, Switzerland.Hepatitis delta virus (HDV) is a viroid-like blood-borne human pathogen that accompanies hepatitis B virus infection in 5% patients. HDV has been studied for four decades; however, the knowledge on its life-cycle and pathogenesis is still sparse. The studies are hampered by the absence of the commercially-available HDV-specific antibodies. Here, we describe a set of reproducible methods for the expression in E. coli of His-tagged small antigen of HDV (S-HDAg), its purification, and production of polyclonal anti-S-HDAg antibodies in rabbits. S-HDAg was cloned into a commercial vector guiding expression of the recombinant proteins with the C-terminal His-tag. We optimized S-HDAg protein purification procedure circumventing a low affinity of the His-tagged S-HDAg to the Ni-nitrilotriacetyl agarose (Ni-NTA-agarose) resin. Optimization allowed us to obtain S-HDAg with >90% purity. S-HDAg was used to immunize Shinchilla grey rabbits which received 80 µg of S-HDAg in two subcutaneous primes in the complete, followed by four 40 µg boosts in incomplete Freunds adjuvant. Rabbits were bled two weeks post each boost. Antibody titers determined by indirect ELISA exceeded 107. Anti-S-HDAg antibodies detected the antigen on Western blots in the amounts of up-to 100 pg. They were also successfully used to characterize the expression of S-HDAg in the eukaryotic cells by immunofluorescent staining/confocal microscopy.publishersversionPeer reviewe

Phosphatidylserine Targets Single-Walled Carbon Nanotubes to Professional Phagocytes In Vitro and In Vivo

Broad applications of single-walled carbon nanotubes (SWCNT) dictate the necessity to better understand their health effects. Poor recognition of non-functionalized SWCNT by phagocytes is prohibitive towards controlling their biological action. We report that SWCNT coating with a phospholipid “eat-me” signal, phosphatidylserine (PS), makes them recognizable in vitro by different phagocytic cells - murine RAW264.7 macrophages, primary monocyte-derived human macrophages, dendritic cells, and rat brain microglia. Macrophage uptake of PS-coated nanotubes was suppressed by the PS-binding protein, Annexin V, and endocytosis inhibitors, and changed the pattern of pro- and anti-inflammatory cytokine secretion. Loading of PS-coated SWCNT with pro-apoptotic cargo (cytochrome c) allowed for the targeted killing of RAW264.7 macrophages. In vivo aspiration of PS-coated SWCNT stimulated their uptake by lung alveolar macrophages in mice. Thus, PS-coating can be utilized for targeted delivery of SWCNT with specified cargoes into professional phagocytes, hence for therapeutic regulation of specific populations of immune-competent cells

Robotic optical telescopes global network MASTER II. Equipment, structure, algorithms

Presented paper describes the basic principles and features of the implementation of a robotic network of optical telescopes MASTER, designed to study the prompt (simultaneous with gamma radiation) optical emission of gamma-ray bursts and to perform the sky survey to detect unknown objects and transient phenomena. With joint efforts of Sternberg astronomical institute, High altitude astronomical station of the Pulkovo observatory, Ural state university, Irkutsk state university, Blagoveshchensk pedagogical university, the robotic telescopes MASTER II near Kislovodsk, Yekaterinburg, Irkutsk and Blagoveshchensk were installed and tested. The network spread over the longitudes is greater than 6 h. A further expansion of the network is considered. © 2011 Springer Science+Business Media B.V

[Methodological recommendations for the use of modern biotechnology for assessing the expression of genes associated with the productivity and resistance of poultry to adverse factors] Методические рекомендации по использованию современных биотехнологий для оценки экспрессии генов, связанных с продуктивностью и устойчивостью птицы к неблагоприятным факторам

Improving the foundations of poultry nutrition is the basis for the dynamic development of agriculture in our country. In this regard, there is no doubt about the importance of studying the microbiota inhabiting the gastrointestinal tract of birds, as well as the mechanisms of activation/inactivation of gene expression, in order to improve methods for assessing metabolism and their health status. Thanks to modern methods, it is possible to assess the effectiveness of the additives on the body, develop schemes for the prevention of diseases and produce high quality poultry products without the use of feed antibiotics, which is the basis for a healthy diet for people. The recommendations are intended for specialists and managers of poultry farms, farmers, researchers, teachers and students of agricultural universities as well as students of the advanced training system. Совершенствование основ питания птиц является базой динамичного развития сельского хозяйства в нашей стране. В связи с этим не вызывает сомнений важность изучения микробиоты, населяющей ЖКТ птиц, а также механизмов активации/инактивации экспрессии генов, с целью совершенствования методов для оценки обмена веществ и состояния их здоровья. Благодаря современным методам удается оценить эффективность действия добавок на организм, разработать схемы профилактики заболеваний и производить птицеводческую продукцию высокого качества без применения кормовых антибиотиков, что является основой здорового питания людей. Рекомендации предназначены для специалистов и руководителей птицеводческих хозяйств, фермеров, научных работников, преподавателей и студентов сельскохозяйственных вузов, слушателей системы повышения квалификации