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21.
Abhilash K Venugopal S Sameer Kumar Ghantasala Lakshmi Dhevi N Selvan Anita Mahadevan Santosh Renuse Praveen Kumar Harsh Pawar Nandini A Sahasrabhuddhe Mooriyath S Suja Yarappa L Ramachandra Thottethodi S Keshava Prasad Shampur N Madhusudhana Harsha HC Raghothama Chaerkady Parthasarathy Satishchandra Akhilesh Pandey Susarla K Shankar 《Clinical proteomics》2013,10(1):3
Introduction
Rabies is a fatal acute viral disease of the central nervous system, which is a serious public health problem in Asian and African countries. Based on the clinical presentation, rabies can be classified into encephalitic (furious) or paralytic (numb) rabies. Early diagnosis of this disease is particularly important as rabies is invariably fatal if adequate post exposure prophylaxis is not administered immediately following the bite.Methods
In this study, we carried out a quantitative proteomic analysis of the human brain tissue from cases of encephalitic and paralytic rabies along with normal human brain tissues using an 8-plex isobaric tags for relative and absolute quantification (iTRAQ) strategy.Results and conclusion
We identified 402 proteins, of which a number of proteins were differentially expressed between encephalitic and paralytic rabies, including several novel proteins. The differentially expressed molecules included karyopherin alpha 4 (KPNA4), which was overexpressed only in paralytic rabies, calcium calmodulin dependent kinase 2 alpha (CAMK2A), which was upregulated in paralytic rabies group and glutamate ammonia ligase (GLUL), which was overexpressed in paralytic as well as encephalitic rabies. We validated two of the upregulated molecules, GLUL and CAMK2A, by dot blot assays and further validated CAMK2A by immunohistochemistry. These molecules need to be further investigated in body fluids such as cerebrospinal fluid in a larger cohort of rabies cases to determine their potential use as antemortem diagnostic biomarkers in rabies. This is the first study to systematically profile clinical subtypes of human rabies using an iTRAQ quantitative proteomics approach. 相似文献22.
Thottethodi Subrahmanya Keshava Prasad Renu Verma Satish Kumar Raja Sekhar Nirujogi Gajanan J Sathe Anil K Madugundu Jyoti Sharma Vinuth N Puttamallesh Anjali Ganjiwale Vithal P Myneedu Aditi Chatterjee Akhilesh Pandey HC Harsha Jayasuryan Narayana 《Clinical proteomics》2013,10(1):8
Background
Purified protein derivative (PPD) has been used for more than half a century as an antigen for the diagnosis of tuberculosis infection based on delayed type hypersensitivity. Although designated as “purified,” in reality, the composition of PPD is highly complex and remains ill-defined. In this report, high resolution mass spectrometry was applied to understand the complexity of its constituent components. A comparative proteomic analysis of various PPD preparations and their functional characterization is likely to help in short-listing the relevant antigens required to prepare a less complex and more potent reagent for diagnostic purposes.Results
Proteomic analysis of Connaught Tuberculin 68 (PPD-CT68), a tuberculin preparation generated from M. tuberculosis, was carried out in this study. PPD-CT68 is the protein component of a commercially available tuberculin preparation, Tubersol, which is used for tuberculin skin testing. Using a high resolution LTQ-Orbitrap Velos mass spectrometer, we identified 265 different proteins. The identified proteins were compared with those identified from PPD M. bovis, PPD M. avium and PPD-S2 from previous mass spectrometry-based studies. In all, 142 proteins were found to be shared between PPD-CT68 and PPD-S2 preparations. Out of the 354 proteins from M. tuberculosis–derived PPDs (i.e. proteins in either PPD-CT68 or PPD-S2), 37 proteins were found to be shared with M. avium PPD and 80 were shared with M. bovis PPD. Alignment of PPD-CT68 proteins with proteins encoded by 24 lung infecting bacteria revealed a number of similar proteins (206 bacterial proteins shared epitopes with 47 PPD-CT68 proteins), which could potentially be involved in causing cross-reactivity. The data have been deposited to the ProteomeXchange with identifier PXD000377.Conclusions
Proteomic and bioinformatics analysis of different PPD preparations revealed commonly and differentially represented proteins. This information could help in delineating the relevant antigens represented in various PPDs, which could further lead to development of a lesser complex and better defined skin test antigen with a higher specificity and sensitivity. 相似文献23.
Michael J Gramer Ewald TJ van den Bremer Muriel D van Kampen Amitava Kundu Peter Kopfmann Eric Etter David Stinehelfer Justin Long Tom Lannom Esther H Noordergraaf Jolanda Gerritsen Aran F Labrijn Janine Schuurman Patrick HC van Berkel Paul WHI Parren 《MABS-AUSTIN》2013,5(6):962-973
The manufacturing of bispecific antibodies can be challenging for a variety of reasons. For example, protein expression problems, stability issues, or the use of non-standard approaches for manufacturing can result in poor yield or poor facility fit. In this paper, we demonstrate the use of standard antibody platforms for large-scale manufacturing of bispecific IgG1 by controlled Fab-arm exchange. Two parental antibodies that each contain a single matched point mutation in the CH3 region were separately expressed in Chinese hamster ovary cells and manufactured at 1000 L scale using a platform fed-batch and purification process that was designed for standard antibody production. The bispecific antibody was generated by mixing the two parental molecules under controlled reducing conditions, resulting in efficient Fab-arm exchange of >95% at kg scale. The reductant was removed via diafiltration, resulting in spontaneous reoxidation of interchain disulfide bonds. Aside from the bispecific nature of the molecule, extensive characterization demonstrated that the IgG1 structural integrity was maintained, including function and stability. These results demonstrate the suitability of this bispecific IgG1 format for commercial-scale manufacturing using standard antibody manufacturing techniques. 相似文献
24.
Liane FM Finotelo Paulo JS Amaral Julio C Pieczarka Edivaldo HC de Oliveira Alcides Pissinati Michaela Neusser Stephan Müller Cleusa Y Nagamachi 《BMC evolutionary biology》2010,10(1):189
Background
The New World monkey (Platyrrhini) subfamily Pitheciinae is represented by the genera Pithecia, Chiropotes and Cacajao. In this work we studied the karyotypes of Pithecia irrorata (2n = 48) and Cacajao calvus rubicundus (2n = 45 in males and 2n = 46 in females) by G- and C-banding, NOR staining and chromosome painting using human and Saguinus oedipus whole chromosome probes. The karyotypes of both species were compared with each other and with Chiropotes utahicki (2n = 54) from the literature. 相似文献25.
Ronald van Eijk Paul HC Eilers Remco Natté Anne-Marie Cleton-Jansen Hans Morreau Tom van Wezel Jan Oosting 《BMC bioinformatics》2010,11(1):67
Background
Multiplex Ligation-Dependent Probe Amplification (MLPA) is an application that can be used for the detection of multiple chromosomal aberrations in a single experiment. In one reaction, up to 50 different genomic sequences can be analysed. For a reliable work-flow, tools are needed for administrative support, data management, normalisation, visualisation, reporting and interpretation. 相似文献26.
Pycnidiospores of Phyllosticta ampelicida, the causal agent of black rot of grape, were found to germinate only on substrata on which they were firmly attached. Such surfaces were poorly wettable and had advancing contact angles (straight thetaa) formed by a water drop of >80°, e.g., grape leaf, polystyrene, Teflon, polycarbonate, collodion, and glass treated with the silanes n-octadecyltrichlorosilane, dimethyldichlorosilane, or diphenyldichlorosilane. When pycnidiospores were deposited on more wettable surfaces they did not attach firmly and did not germinate. Such highly wettable surfaces had straight thetaa = 40° and were represented by heat-treated glass, cellophane, nutrient- and water-agars, polystyrene treated with UV-irradiation or sulfuric acid, and glass silanized with n-2-aminoethyl-3-aminopropyltrimethoxysilane, n-(trimethoxysilylpropyl)ethylenediamine triacetic acid trisodium, or 3-aminopropyltriethoxysilane. Adhesion of pycnidiospores was assessed with and without a hydraulic shearing force. Pycnidiospore adhesion occurred over several minutes in distilled deionized water, unless it was first acidified, which decreased attachment time to <0.03 s. Attachment of pycnidiospores treated with sodium azide, formaldehyde, or boiled in water for 10 min was similar to nontreated conidia. Possible mechanisms of adhesion of the conidia to surfaces include hydrophobic and ionic interactions. 相似文献