Fluorescence polarization assay, competitive enzyme-linked immunosorbent assay (ELISA-C) and indirect ELISA for the diagnosis of brucellosis in buffaloes (Bubalus bubalis)

The objective of the present study was to compare the performance of three serological tests for diagnosis of Brucella abortus infections in buffaloes (Bubalus bubalis). Serum samples collected from 696 adult females were submitted to the competitive enzyme-linked immunosorbent assay (ELISAC), (I-ELISA), fluorescence polarization test (FPA), 2-mercaptoethanol test (2-ME) and complement fixation test (CFT). The gold standard was the combination of CFT and 2-ME, considering as positive the reactors in both CFT and 2-ME, and as negative those non-reactors. ROC analyses were done for C-ELISA, I-ELISA and FPA and the Kappa agreement index were also calculated. The best combinations of relative sensitivity (SEr) and relative specificity (SPr) and Kappa were given by C-ELISA (96.9%, 99.1%, and 0.932, respectively) and FPA (92.2%, 97.6 and 0.836, respectively). The C-ELISA and FPA were the most promising confirmatory tests for the serological diagnosis of brucellosis in buffaloes, and for these tests, cut-off values for buffaloes may be the same as those used for bovines.

Discrepancies and consequences of indirect hemagglutination, indirect immunofluorescence and Elisa tests for the diagnosis of Chagas disease

Using the indirect hemagglutination (IH), indirect immunofluorescence (IIF) and enzyme linked immunosorbent assay (ELISA) tests for the diagnosis of Chagas disease, 4000 serum samples were examined. This study was conducted with different purposes: clinical interest, research support and parasitological monitoring of those patients with Chagas disease who were treated with heart transplantations. The tests occurred without patient selection and in accordance with the medical requests. The results showed discrepancies and brought about several questions, considering the different results that all three methods showed when considered together. What was found brought about concerns and we suggest the adoption of different measures, aiming to avoid these mismatches in the context of this disease.

Characterization of NiCrAlC PTA coatings

Surface tailoring with plasma transferred arc (PTA) hardfacing involves the deposition of powder mixtures to produce coatings with an almost unlimited chemical composition. PTA hardfacing is particularly important for processing low weldability alloys, such as those for high-temperature applications, of which NiCrAlC is an example. This study analyzed NiCrAlC coatings processed by PTA using a mixture of elemental powders. Deposition on AISI316L plates was carried out with currents of 100 A and 130 A to induce variations in the chemical composition of the coatings, which were also subjected to isothermal exposure at temperatures of up to 1000 ºC for up to 72 hours in an air furnace. The results show that the aluminide compounds were stable, the coatings that solidified more slowly having the highest hardness after temperature exposure and potentially being suitable for use in high-temperature abrasion environments.

Sviluppo di un modello quantitativo di valutazione e valorizzazione del servizio di taglio materiali: il caso G.D

La necessità di sviluppare questo progetto è nata dal fatto che quando due aziende desiderano collaborare in maniera costruttiva con rapporti di fornitura di materiali o lavorazioni, bisogna evitare che si verifichino scostamenti nei rapporti a discapito di uno dei due contraenti. La coscienza dell'inefficienza di un rapporto squilibrato ha portato le due aziende a intavolare un progetto migliorativo basato sulla massima trasparenza e propositività. Quando G.D ha deciso di avvalersi della fornitura da parte di Transmec per quanto riguarda il servizio di taglio del materiale, l'ha fatto per andare a cercare un miglior livello di efficacia ed efficienza. Il raggiungimento degli attuali livelli di performance non sarebbe stato possibile se entrambe le aziende non avessero collaborato attivamente nell'ottenimento degli standard desiderati. Questo studio è nato dal fatto che il modello interno adottato precedentemente da G.D per la ripartizione dei costi delle Business Unit non era più efficace se utilizzato come strumento di pagamento. Si rendeva quindi necessaria un'analisi accurata della situazione produttiva utilizzando gli approcci analitici resi disponibili nel corso degli anni come lo strumento per l'allocazione dei costi secondo le attività (Activity Based Costing) e la mappatura dei processi utilizzata nei casi di re-ingegnerizzazione dei reparti.

PTA e stenting carotideo: valutazione morfologica ed evolutiva, a breve e medio termine

In un periodo di tre anni è stato svolto un lavoro mirato alla valutazione delle complicanze correlate all’utilizzo dello stenting carotideo. Dopo la preparazione di un protocollo con definizione di tutti i fattori di rischio sono stati individuati i criteri di inclusione ed esclusione attraverso i quali arruolare i pazienti. Da Luglio 2004 a Marzo 2007 sono stati inclusi 298 pazienti e sono state valutate le caratteristiche della placca carotidea, con particolare riferimento alla presenza di ulcerazione e/o di stenosi serrata, la tortuosità dei vasi e il tipo di arco aortico oltre a tutti i fattori di rischio demografici e metabolici. E’ stato valutato quanto e se questi fattori di rischio incrementino la percentuale di complicanze della procedura di stenting carotideo. I pazienti arruolati sono stati suddivisi in due gruppi a seconda della morfologia della placca: placca complicata (placca con ulcera del diametro > di 2 mm e placca con stenosi sub occlusiva 99%) e placca non complicata. I due gruppi sono stati comparati in termini di epidemiologia, sintomatologia neurologica preoperatoria, tipo di arco, presenza di stenosi o ostruzione della carotide controlaterale, tipo di stent e di protezione cerebrale utilizzati, evoluzione clinica e risultati tecnici. I dati sono stati valutati mediante analisi statistica di regressione logistica multipla per evidenziare le variabili correlate con l’insuccesso. Dei 298 pazienti consecutivi sottoposti a stenting, 77 hanno mostrato una placca complicata (25,8%) e 221 una placca non complicata (74,2%). I due gruppi non hanno avuto sostanziali differenze epidemiologiche o di sintomatologia preoperatoria. Il successo tecnico si è avuto in 272 casi (91,2%) e sintomi neurologici post-operatosi si sono verificati in 23 casi (23.3%). Tutti i sintomi sono stati temporanei. Non si sono avute differenze statisticamente significative tra i due gruppi in relazione alle complicanze neurologiche e ai fallimenti tecnici. L’età avanzata è correlata ad un incremento dei fallimenti tecnici. I risultati dello studio portano alla conclusione che la morfologia della placca non porta ad un incremento significativo dei rischi correlati alla procedura di stenting carotideo e che l’indicazione alla CAS può essere posta indipendentemente dalla caratteristica della placca.

Emotional design e genetica dello stile in campo automotive: studio qualitativo e quantitativo delle linee di carattere

La figura del designer detiene un ruolo chiave nella progettazione di un nuovo prodotto: a lui spetta l'importante compromesso fra scelte ingegneristiche, di mercato e di stile. Lo studio qui condotto analizza da un punto di vista qualitativo e quantitativo le linee di carattere che delineano il profilo base di un'autovettura, in riferimento al particolare segmento automobilistico ed al suo stile. In tal modo si vuole porre una base alla "genetica dello stile", ovvero la caratterizzazione matematica dello stile.

Characterization of antibodies specific for canine TLR4, 5 and 9 by ELISA, Western blotting and immunohistochemistry

Toll-like receptors recognize pathogen-associated molecular patterns of microbial origin, and ligand recognition results in the production of different immune mediators such as pro-inflammatory cytokines, interferon, reactive oxygen and nitrogen intermediates, and upregulation of costimmulatory molecules. As these receptors have a critical role in linking pathogen recognition to induction of inflammation and innate as well as adaptive immunity, there is tremendous interest in understanding how the tissue and cell-type expression of TLRs is regulated and its influence on the local innate immune response. While TLRs are well studied in humans and rodents, to date little is known about them in dogs. The purpose of this study was to develop canine specific antibodies against TLR2, 4, 5 and 9 that were used to measure relative expression of these TLRs in healthy and reactive canine mesenteric lymph nodes. All 8 rabbit sera (2 each for TLR2, 4, 5 and 9) were strongly positive in ELISA against the respective 2 peptides per TLR used for immunization. The purified antibodies selected specifically detected a protein band with an apparent size of approximately 70 kDa in lysates of canine PBMCs by Western blotting. Immunostaining was observed with purified antibodies against TLR4, 5 and 9, whereas for canine TLR2, staining was only observed with the unpurified antibodies. In the mesenteric lymph node of healthy dogs, the overall staining pattern was very similar for TLR4 and 5 with positive cells predominantly found in the internodular areas and lower part of the cortex. Compared to the TLR4 and 5, more cells stained positive for TLR9 especially in the lymphoid nodules. The reactive lymph nodes contained more TLR4 and 9 positive cells. Moreover, a shift of TLR-9 positive cells from the lymphoid follicles to the deep cortex and medullary cords was observed. Whereas TLR9 co-localized with CD79-positive areas, TLR4 and 5 antibodies stained cells primarily in the CD3-positive areas. All three TLR antibodies stained cells within the area that co-localized with lysozyme-positive cells. In conclusion, this study demonstrates that the antibodies generated against canine TLR 4, 5 and 9 identify the expression of these TLRs in formalin-fixed canine lymph nodes and demonstrate increased expression in reactive canine mesenteric lymph nodes.

Post-surgical follow-up (by ELISA and immunoblotting) of cured versus non-cured cystic echinococcosis in young patients

The study was designed to determine comparatively the prognostic value of immunoblotting and ELISA in the serological follow-up of young cystic echinococcosis (CE) patients exhibiting either a cured or a progredient (non-cured) course of disease after treatment. A total of 54 patients (mean age 9 years, range from 3 to 15 years) with surgically, radiologically and/or histologically proven CE were studied for a period up to 60 months after surgery. Additionally, some of the patients underwent chemotherapy. Based on the clinical course and outcome, as well as on imaging findings, patients were clustered into 2 groups of either cured (CCE), or non-cured (NCCE) CE patients. ELISA showed a high rate of seropositivity 4 to 5 years post-surgery for both CCE (57.1%) and NCCE (100%) patients, the difference found between the two groups was statistically not significant. Immunoblotting based upon recognition of AgB subcomponents (8 and 16 kDa bands) showed a decrease of respective antibody reactivities after 4 years post-surgery. Only sera from 14.3% of CCE patients recognized the subcomponents of AgB after 4 years, while none (0%) of these sera was still reactive at 5 years post-surgery. At variance, immunoblotting remained positive for AgB subcomponents in 100% of the NCCE cases as tested between 4 and 5 years after surgical treatment. Immunoblotting therefore proved to be a useful approach for monitoring post-surgical follow-ups of human CCE and NCCE in young patients when based upon the recognition of AgB subcomponents.

Sensitive and highly specific quantitative real-time PCR and ELISA for recording a potential transfer of novel DNA and Cry1Ab protein from feed into bovine milk

To address food safety concerns of the public regarding the potential transfer of recombinant DNA (cry1Ab) and protein (Cry1Ab) into the milk of cows fed genetically modified maize (MON810), a highly specific and sensitive quantitative real-time PCR (qPCR) and an ELISA were developed for monitoring suspicious presence of novel DNA and Cry1Ab protein in bovine milk. The developed assays were validated according to the assay validation criteria specified in the European Commission Decision 2002/657/EC. The detection limit and detection capability of the qPCR and ELISA were 100 copies of cry1Ab microL(-1) milk and 0.4 ng mL(-1) Cry1Ab, respectively. Recovery rates of 84.9% (DNA) and 97% (protein) and low (<15%) imprecision revealed the reliable and accurate estimations. A specific qPCR amplification and use of a specific antibody in ELISA ascertained the high specificity of the assays. Using these assays for 90 milk samples collected from cows fed either transgenic (n = 8) or non-transgenic (n = 7) rations for 6 months, neither cry1Ab nor Cry1Ab protein were detected in any analyzed sample at the assay detection limits.

Application of Immunohistochemistry and ELISA for the Diagnosis of Neospora-Infected Cattle

Studies were undertaken to adapt diagnostic methods for use in our laboratory for detection of Neospora sp. infection in cattle. An immunohistochemical (IHC) test was used for detection of Neospora sp. antigen in tissues of aborted bovine fetuses. Neospora sp. antigen was detected most frequently in fetal brain tissue. Polyclonal antibodies were tested for specificity and sensitivity of the IHC. Sera were obtained from Neospora sp. infected dairy herds for use as positive and negative controls in the continuing development of an enzyme-linked immunosorbent assay (ELISA).