PMab-219: A monoclonal antibody for the immunohistochemical analysis of horse podoplanin

Monoclonal antibodies (mAbs) against human, mouse, rat, rabbit, dog, cat, and bovine podoplanin (PDPN), a lymphatic endothelial cell marker, have been established in our previous studies. However, mAbs against horse PDPN (horPDPN), which are useful for immunohistochemical analysis, remain to be developed. In the present study, mice were immunized with horPDPN-overexpressing Chinese hamster ovary (CHO)-K1 cells (CHO/horPDPN), and hybridomas producing mAbs against horPDPN were screened using flow cytometry. One of the mAbs, PMab-219 (IgG_2a, kappa), specifically detected CHO/horPDPN cells via flow cytometry and recognized horPDPN protein using Western blotting. Furthermore, PMab-219 strongly stained CHO/horPDPN via immunohistochemistry. These findings suggest that PMab-219 is useful for investigating the function of horPDPN.     

Evaluation of the allergenicity of ω5-gliadin-deficient Hokushin wheat (1BS-18) in a wheat allergy rat model

We previously developed Hokushin wheat line as a hypoallergenic wheat lacking ω5-gliadin (1BS-18), a major allergen for wheat-dependent exercise-induced anaphylaxis. However, the allergenicity of 1BS-18 has not been understood completely. In this study, we evaluated the allergenicity of 1BS-18 such as anaphylactic elicitation ability and sensitization ability using rats sensitized with ω5-gliadin or glutens prepared from Hokushin (Hokushin gluten) or 1BS-18 (1BS-18 gluten). Rats were sensitized by intraperitoneal administration of ω5-gliadin, Hokushin gluten or 1BS-18 gluten. Immunoglobulin E-mediated systemic anaphylaxis was evaluated by measuring changes in rectal temperature for 30 min after intravenous challenge with ω5-gliadin or the test glutens in unsensitized rats or rats sensitized with ω5-gliadin or the test glutens. In ω5-gliadin-sensitized rats, intravenous challenge with ω5-gliadin or Hokushin gluten significantly decreased the rectal temperature at 30 min after challenge while challenge with 1BS-18 gluten did not reduce the rectal temperature. Furthermore, intravenous challenge with ω5-gliadin significantly decreased the rectal temperature in rats sensitized with Hokushin gluten or 1BS-18 gluten. However, the reduced degree observed in 1BS-18 gluten-sensitized rats was smaller than that in Hokushin gluten-sensitized rats. In conclusion, 1BS-18 elicited no allergic reaction in ω5-gliadin-sensitized rats and had less sensitization ability for ω5-gliadin than that of Hokushin wheat.     

Ratiometric assay of CARM1 activity using a FRET-based fluorescent probe

One of the regulatory mechanisms of epigenetic gene expression is the post-translational methylation of arginine residues, which is catalyzed by protein arginine methyltransferases (PRMTs). Abnormal expression of PRMT4/CARM1, one of the PRMTs, is associated with various diseases, including cancers. Here, we designed and synthesized a Förster resonance energy transfer (FRET)-based probe, FRC, which contains coumarin and fluorescein fluorophores at the N-terminus and C-terminus of a peptide containing an arginine residue within an appropriate amino acid sequence to serve as a substrate of CARM1; the two fluorophores act as a FRET donor and a FRET acceptor, respectively. Since trypsin specifically hydrolyzes the arginine residue, but not a monomethylarginine or dimethylarginine residue, CARM1 activity can be evaluated from the change of the coumarin/fluorescein fluorescence ratio of FRC in the presence of trypsin.     

Wie V?gel ihr Auge schützen: Zur Arbeitsteilung von Oberlid, Unterlid und Nickhaut

Zusammenfassung Vögel schließen ihre Augen im Schlaf in einer für die großen Taxa typischen Weise. Entweder geht das Unterlid hoch wie bei der Mehrzahl der Arten, oder das Oberlid bewegt sich abwärts (Psittaciformes, Trochili), oder aber beide Lider schließen die Lidspalte (Strigiformes, Caprimulgi). Solche Kenntnis fehlt von den meisten Ordnungen, oder die Handbücher geben falsche oder widersprüchliche Information. Neben dem tonischen, schlafbegleitenden Augenschluss bewegen Vögel im Wachzustand eines oder beide Lider phasisch und meist schnell. Dieser häufige Lidschlag ist durch ein anderes Bewegungsmuster und durch eine andere Funktion gekennzeichnet. Photodokumente und genaue Beobachtungen führen erstmals zu einer funktionellen Deutung, der zufolge der Lidschlag das Auge mechanisch schützt. Droht dem Auge von vorn oder von oben eine potentielle Schädigung, so schließt das Oberlid bei Tauben, Eulen und Singvögeln, im Sprühwasser gleichzeitig auch das Unterlid (Cinclus). Der unabweisbarste Beleg stammt aus dem Vergleich des Aufpickens dorniger, sperriger Beuteinsekten mit Oberlidschluss gegenüber dem Aufnehmen harmloser Beeren ohne jede Lidbewegung (Gallicolumba). Weiter ist die Antwort des Oberlids, anders als beim Unterlid, öfter seitengerecht reizorientiert, so dass die Bewegung einseitig sein kann. Zudem kann der Schluss des Oberlids auch bei stationärem (Feind-)reiz seitenweise alternieren (Otus). Ausnahmsweise tritt eine adaptive Asymmetrie auch während kurzer Zeiten der Augenöffnung zum Spähen nach Feinden im Schlaf auf, und zwar hier beim Unterlid der bedrohten Seite (Anas).Eine neue Funktion wird auch dem Schlag der Nickhaut (Membrana nictitans) zugeschrieben. Traditionell als die Cornea reinhaltendes Organ gesehen, dient auch sie dem mechanischen Schutz des Auges. Auch sie kann seitenrichtig reizorientiert schlagen, doch ist hierüber wenig bekannt. Dieselben Reize, die den Lidschlag auslösen, können bei anderen Arten die Nickhaut schlagen lassen. Ihre Schlagrate ist schwierig zu messen, da viele Schläge (nur?) mit denen des Oberlides zusammenfallen und so verborgen bleiben (Otus). Diese Synchronie ist mit keiner der bisher vorgeschlagenen Funktionen erklärbar, ebenso wenig wie die verborgenen Schläge bei tonischem Augenschluss (Passer).Die Annahme einer Ausschaltung störender Sinnesinformation, z.B. während rascher Kopfbewegungen, durch die Nickhaut lässt sich aus vier Gründen verwerfen. Die Zunahme der Schlagrate während des Feindalarms (Ficedula) bleibt funktionell unerklärt.In einer bei Vögeln einzigartigen Weise schützt der Samtkleiber (Sitta azurea) sein Auge durch Zusammenziehen des nackten Augenrings (Lidblende), wenn er rücklings an der Unterseite von Ästen nahrungssuchend einem ständigen Regen von losgelösten Rindenteilchen u. ä. ausgesetzt ist.Sekundär haben sich die Bewegungen eines oder beider Lider oder aber der Nickhaut zu optischen Signalen entwickelt, und zwar durch kontrastierende Feder- oder Nickhautfärbung. Die betreffenden blitzschnell aufleuchtenden Signale sind an den Paarpartner (Cinclus, Corvidae,Cepphus), an mögliche Feinde (Anas) oder an bisher unbekannte Empfänger gesichtet (Ficedula).

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On how birds protect their eyes: division of labour between the upper lid, lower lid and the nictitating membrane

Summary Birds close their eyes during sleep in various taxon-specific ways. Either the lower lid moves up as in the majority of species including the Anseres, Accipitres, Falconiformes, Galli, Charadrioidea, Columbiformes, and Oscines; or the upper lid moves down (Psittaciformes, Trochili), or both lids close the eye as in Strigiformes and Caprimulgi. Such information is absent for most orders, or the handbooks provide wrong or conflicting information. Beside the tonic, sleep-related eye closure, birds move one or both lids in a phasic, usually swift mode when awake. These frequent lid movements are typified by their different co-ordination and function. Photographic and observational evidence strongly suggests mechanical protection of the eye as a novel function (where this had not been proposed previously). When an impact from any object is imminent from in front of or above the head, the upper lid shuts in pigeons, owls and oscines, and with water splashing, the lower lid as well (Cinclus). The most convincing evidence for mechanical protection comes from the deployment of the upper lid during the picking up of spiny insect prey as compared to easy-to-swallow berries, when both lids stay at rest (Gallicolumba).Further, the response of the upper lid is more stimulus-oriented so that both upper lids move asymmetrically. But there is also a unilateral, alternating winking of the upper lids when causative (predator) stimuli remain stationary. This never occurs with the lower lids (Otus). As an exception, an adaptive asymmetry occurs during brief phases of unilateral scanning interrupting sleep, designed to detect approaching predators. This scanning involves the lower lid (Anas).A new function is also attributed to the beating of the nictitating membrane (Membrana nictitans). Traditionally viewed as a cleaning device it also serves to protect the eye from mechanical impact, and it also can be tuned to the side from where danger is threatening, though by and large there is a dearth of information from avian taxa. The non-visually elicited action of the membrane seems always to be bilateral (Falco, Harpia). The very stimuli eliciting the blinking of a lid can, in different species, trigger the beat of the membrane, and can cause it to move tonically (Falco). The membrane beats at a rate difficult to measure since many of its beats coincide with the blinking of the upper lid and thus remain hidden (Otus). This coincidence is difficult to account for by any function discussed so far, as are the many hidden beats during tonic eye closure with the lids (Passer).The hypothesis according to which the action of the membrane is filtering out undesirable retinal stimulation during e.g. rapid head movements is dismissed on four different grounds. The increase of the membrane activity during predator alarm (Ficedula) is functionally unaccounted for.In a fashion unique among birds, the Blue Nuthatch (Sitta azurea) protects its eyes by contracting the naked skin surrounding the eye, thereby minimizing the exposure of the cornea; during foraging along the underside of branches, a continual rain of bark particles and debris jeopardizes unimpeded vision.Secondarily, one or both lids or the nictitating membrane have taken on the function of optic signals by virtue of contrasting feather colour or coloration. The phasic (flashing) signal movements involved are directed at the pair mate (Cinclus, Corvidae,Cepphus), predators (Anas) or at unknown parties (Ficedula).

Dies ist Veröffentlichung Nr. 29 des Philippine Endemic Species Conservation Project der Zoologischen Gesellschaft Frankfurt.     

Androgen receptor complex binding in murine skeletal muscle nuclei

Examination of binding of androgen-receptor complexes from murine skeletal muscle cytosol was performed by modified nuclear retention assay and modified nuclear acceptor assay. The experiments showed the binding of androgen-receptor complexes to the nuclear acceptor sites to be a cooperative process. Hill analysis of the data obtained resulted in a Hill coefficient of 3,6. The apparent dissociation constant for binding of cytosolic [³H]-testosterone-receptor complexes to nuclei was found to be in the range of K_D = 6 ? 8 × 10^?11 M. The nuclear matrix was able to bind androgen-receptor complexes in a saturable way, too.     

中国海洋渔业水域环境研究现状及展望

渔业水域环境是水生生物赖以生存、繁衍的最基本条件,优良的渔业水域环境是海洋渔业持续发展的物质载体与必然条件。近年来中国海洋渔业水域环境状况日趋严峻,依据海洋渔业水域监测数据,分析了中国海洋渔业水域环境的现状;概括了中国海洋渔业水域环境面临的主要问题及破坏原因;综述了海洋渔业水域环境保护及修复对策的研究现状,并对今后的研究工作提出了方向与内容上的展望。     

Non-invasive measurement of small peptides in the common marmoset (Callithrix jacchus): a radiolabeled clearance study and endogenous excretion under varying social conditions

A non-invasive assay for measurement of oxytocin (OT) and vasopressin (AVP) in primates would enable researchers to study the relationship between the endocrine system and behavior without disturbing potentially endangered animals in their natural habitats. In order to test whether or not OT specifically would be measurable in the urine of a primate, 10 microCi of tritium-labeled OT were injected into the peripheral blood supply of four adult male common marmosets (Callithrix jacchus), with continuous urinary collection over 48 h. When urine was processed by HPLC separation and beta counting for radioactive clearance, the label was present in all samples in the fraction where OT elutes. Large amounts of OT were also seen in a fraction other than that containing the OT standard, indicating that OT is measurable but that it also undergoes substantial metabolic breakdown. In a second experiment, we isolated six common marmosets for 48 h and then exposed them to social contact to evaluate the effect of changing social stimuli on endogenous urinary measurement of both OT and AVP. Both were measured after HPLC separation to isolate the intact molecule and also to control for cross-reactivity with metabolites in subsequent RIA. Cortisol was also measured to objectively evaluate the stress response. A priori assumptions were that urinary OT and AVP would be lower during a period of isolation and higher during periods of social contact. These assumptions were met, leading us to conclude that peripheral OT and AVP are measurable via urinary assay and that such an assay is a valid means of evaluating social condition in this species.     

Barnase-barstar high affinity interaction phenomenon as the base for the heterogenous bioluminescence pseudorabies virus' immunoassay

The effective new variant of "sandwich" bioluminescent enzyme immunoassay (BEIA) for the sensitive detection of glycoprotein B (gB) of pseudorabies virus (PrV) was presently developed. The high affinity interaction of barnase-barstar protein pair and photoprotein obelin as bioluminescent marker were for the first time successfully applied to BEIA development. Preliminary the two monoclonal antibodies, 11/5 and 34/2, were raised against gB for ELISA PrV detection. Presently we used the same immuno-"sandwich" principle for BEIA. To do this the two different bioconjugates were elaborated. Recombinant barnase was chemically conjugated with monoclonal anti-PrV's gB IgG, and also barstar was fused in frame to obelin. The characteristics of BEIA method have been compared to ELISA PrV detection. We have shown the proposed here gB-BEIA was 40-fold more sensitive as opposed to gB-ELISA test. The construction might have a broad promise in multiple potential immunological applications.