Getting hold of the tobamovirus particle : why and how? Purification routes over time and a new customizable approach

This article develops a multi-perspective view on motivations and methods for tobamovirus purification through the ages and presents a novel, efficient, easy-to-use approach that can be well-adapted to different species of native and functionalized virions. We survey the various driving forces prompting researchers to enrich tobamoviruses, from the search for the causative agents of mosaic diseases in plants to their increasing recognition as versatile nanocarriers in biomedical and engineering applications. The best practices and rarely applied options for the serial processing steps required for successful isolation of tobamoviruses are then reviewed. Adaptations for distinct particle species, pitfalls, and ‘forgotten’ or underrepresented technologies are considered as well. The article is topped off with our own development of a method for virion preparation, rooted in historical protocols. It combines selective re-solubilization of polyethylene glycol (PEG) virion raw precipitates with density step gradient centrifugation in biocompatible iodixanol formulations, yielding ready-to-use particle suspensions. This newly established protocol and some considerations for perhaps worthwhile further developments could serve as putative stepping stones towards preparation procedures appropriate for routine practical uses of these multivalent soft-matter nanorods.Part of this work was funded by the Deutsche Forschungsgemeinschaft (DFG: German Research Foundation), project number 446507449.Deutsche Forschungsgemeinschaft (DFG: German Research Foundation

Facile purification and use of tobamoviral nanocarriers for antibody-mediated display of a two-enzyme system

Immunosorbent turnip vein clearing virus (TVCV) particles displaying the IgG-binding domains D and E of Staphylococcus aureus protein A (PA) on every coat protein (CP) subunit (TVCVPA) were purified from plants via optimized and new protocols. The latter used polyethylene glycol (PEG) raw precipitates, from which virions were selectively re-solubilized in reverse PEG concentration gradients. This procedure improved the integrity of both TVCVPA and the wild-type subgroup 3 tobamovirus. TVCVPA could be loaded with more than 500 IgGs per virion, which mediated the immunocapture of fluorescent dyes, GFP, and active enzymes. Bi-enzyme ensembles of cooperating glucose oxidase and horseradish peroxidase were tethered together on the TVCVPA carriers via a single antibody type, with one enzyme conjugated chemically to its Fc region, and the other one bound as a target, yielding synthetic multi-enzyme complexes. In microtiter plates, the TVCVPA-displayed sugar-sensing system possessed a considerably increased reusability upon repeated testing, compared to the IgG-bound enzyme pair in the absence of the virus. A high coverage of the viral adapters was also achieved on Ta2O5 sensor chip surfaces coated with a polyelectrolyte interlayer, as a prerequisite for durable TVCVPA-assisted electrochemical biosensing via modularly IgG-assembled sensor enzymes.Deutsche Forschungsgemeinschaft (DFG: German Research Foundation)Nomad Bioscience GmbHGerman Research Foundation (DFG

History as a rhetorical instrument in Tertullian's Ad Nationes : a critical investigation

Thesis (MA (Latin))--North-West University, Potchefstroom Campus, 2012This study traced Tertullian’s utilisation of history (or historical material) as a rhetorical instrument in one of his earliest works, the Ad Nationes. An in-depth analysis of the book identified this as a fundamental trajectory in the argument of Tertullian. The study casts a new perspective on the written work of this renowned Christian apologist and theologian. His use of history particularly to substantiate his arguments was compared with the contemporary primary sources, in order to assess the integrity or accuracy of his historical data. The prevailing rhetoric, as e.g. outlined by Quintilian, valued the message and intention of a text higher than the historical accuracy of the account. The same Quintilian, however, emphasized that historical accuracy would guarantee the message and intention of a text. The research concluded that Tertullian, who enjoyed a classical education and was therefore well acquainted with the rules of rhetoric, did pay sufficient attention to Quintilian’s insistence on historical accuracy in his utilisation of history. Tertullian was well aware of the significance of historical accuracy. On occasion he rightly criticised Tacitus (the famous historian) for historical inaccuracies in his work. In his Apologeticus (in which much of the Ad Nationes was reworked) he corrected some historical data. In the Ad Nationes he wrote a brilliant paragraph on the origin of rumours (fama) and also expressed his appreciation for careful investigation (in court procedures) in order to ascertain the truth (veritas) accurately. In the rhetorical utilisation of historical material, accurate historical knowledge did not play a crucial role. Of paramount importance was the intention and purpose of the immediate argument.Master

Five brave pagan women in the work of Tertullian

Historical exempla have been employed as a rhetorical stratagem since the 8th century BCE. The Christian apologist Tertullian, who received thorough training in rhetoric, was familiar with this device. Lucretia, Cleopatra, Dido, ‘the wife of Hasdrubal’ and Leaena also appear in the works of Roman authors as typical examples of bravery. In his Ad Martyres (4), Tertullian groups together all five as fine examples of bravery when facing death. Four of the examples appear in his Ad Nationes (1.18) and two in the Liber Apologeticus (50). Both the Ad Nationes and the Liber Apologeticus were written in defence of the Christians. In Tertullian’s De exhortatione castitatis (13.3) and the De Monogamia (17), Lucretia and Dido are referred to as exempla of chastity and monogamy. The first four paragraphs preceding the analysis of these texts provide the context and contain background information on the Christian apologists, a short biography of Tertullian,a few remarks on the use of history as a rhetorical tool and a discussion on the place of these women in ancient pagan literature. The investigation of Tertullian’s use of the exempla to bolster his argument reveals how he adapted these stock examples of pagan women to fit the Christian context, and illustrates how and why ancient rhetoric became part of Christian literary tradition. Vyf dapper heidense vroue in die werk van Tertullianus. Historiese exempla word al sedertdie agtste eeu v.C. as ’n retoriese strategie benut. Die Christen apologeet, Tertullianus, wat ’n grondige opleiding in retoriek ontvang het, was vertroud met hierdie tegniek. Lucretia, Cleopatra, Dido, ‘Hasdrubal se vrou’ en Leaena kom ook in die werke van Romeinse outeurs voor as tipiese voorbeelde van dapperheid. In sy Ad Martyres (4) groepeer Tertullianus al vyf saam as goeie voorbeelde van dapperheid in die aangesig van die dood. Vier van die voorbeelde kom in die Ad Nationes (1.18) voor en twee in sy Liber Apologeticus (50). Albei hierdie werke is geskryf om die Christene te verdedig. In Tertullianus se De exhortatione castitatis (13.3) en die De Monogamia (17) word na Lucretia en Dido as voorbeelde van kuisheid en monogamie verwys. Die eerste vier paragrawe wat die analise voorafgaan, gee die konteks en bevat agtergrondinligting oor die Christen apologete, ’n kort biografie van Tertullianus, ’n paar opmerkings oor die gebruik van geskiedenis as retoriese instrument en ’n bespreking van die plek van die vyf vroue in antieke heidense letterkunde. Die ondersoek na Tertullianus se gebruik van hierdie exempla om sy argument te versterk, dui aan dat hy die standaardvoorbeelde van heidense vroue aangepas het om by die Christelike konteks te pas, en illustreer hoe en waarom antieke retoriek deel van die Christelike literêre tradisie geword het

Getting Hold of the Tobamovirus Particle—Why and How? Purification Routes over Time and a New Customizable Approach

This article develops a multi-perspective view on motivations and methods for tobamovirus purification through the ages and presents a novel, efficient, easy-to-use approach that can be well-adapted to different species of native and functionalized virions. We survey the various driving forces prompting researchers to enrich tobamoviruses, from the search for the causative agents of mosaic diseases in plants to their increasing recognition as versatile nanocarriers in biomedical and engineering applications. The best practices and rarely applied options for the serial processing steps required for successful isolation of tobamoviruses are then reviewed. Adaptations for distinct particle species, pitfalls, and ‘forgotten’ or underrepresented technologies are considered as well. The article is topped off with our own development of a method for virion preparation, rooted in historical protocols. It combines selective re-solubilization of polyethylene glycol (PEG) virion raw precipitates with density step gradient centrifugation in biocompatible iodixanol formulations, yielding ready-to-use particle suspensions. This newly established protocol and some considerations for perhaps worthwhile further developments could serve as putative stepping stones towards preparation procedures appropriate for routine practical uses of these multivalent soft-matter nanorods

Facile Purification and Use of Tobamoviral Nanocarriers for Antibody-Mediated Display of a Two-Enzyme System

Immunosorbent turnip vein clearing virus (TVCV) particles displaying the IgG-binding domains D and E of Staphylococcus aureus protein A (PA) on every coat protein (CP) subunit (TVCVPA) were purified from plants via optimized and new protocols. The latter used polyethylene glycol (PEG) raw precipitates, from which virions were selectively re-solubilized in reverse PEG concentration gradients. This procedure improved the integrity of both TVCVPA and the wild-type subgroup 3 tobamovirus. TVCVPA could be loaded with more than 500 IgGs per virion, which mediated the immunocapture of fluorescent dyes, GFP, and active enzymes. Bi-enzyme ensembles of cooperating glucose oxidase and horseradish peroxidase were tethered together on the TVCVPA carriers via a single antibody type, with one enzyme conjugated chemically to its Fc region, and the other one bound as a target, yielding synthetic multi-enzyme complexes. In microtiter plates, the TVCVPA-displayed sugar-sensing system possessed a considerably increased reusability upon repeated testing, compared to the IgG-bound enzyme pair in the absence of the virus. A high coverage of the viral adapters was also achieved on Ta2O5 sensor chip surfaces coated with a polyelectrolyte interlayer, as a prerequisite for durable TVCVPA-assisted electrochemical biosensing via modularly IgG-assembled sensor enzymes