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Gary D. Pipes Phillip Campbell Pavel V. Bondarenko Bruce A. Kerwin Michael J. Treuheit Himanshu S. Gadgil 《Journal of pharmaceutical sciences》2010,99(11):4469-4476
A middle-down LC/MS approach, for the rapid quantitation and characterization of site-specific methionine oxidation in a recombinant monoclonal IgG1 molecule, is described. An IgG1 antibody was digested with endoprotease LysC under limited proteolytic conditions to produce two major components; an antigen binding fragment (Fab) and a crystallizable fraction (Fc). These fractions were then reduced to produce three major species; light chain (LC), Fc/2 which is the C terminal region of the heavy chain (HC) and the N-terminal heavy chain region (Fd). These three fragments were separated by reversed-phase HPLC using a diphenyl column. The diphenyl column resolved site-specific methionine oxidation in all three subunits. Middle- down N-terminal sequencing with a LCT premier mass spectrometer was used to identify the sites of oxidation in the LC. Sites of oxidation in the Fc/2 were identified using middle-down collision-induced dissociation (CID) on a Qtof premier. This method allowed for the rapid quantitation and identification of oxidation on each methionine residue in an IgG1 molecule. 相似文献
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A tumor-targeted and conditionally replicating oncolytic adenovirus vector expressing TRAIL for treatment of liver metastases. 总被引:3,自引:0,他引:3
Pavel Sova Xiao-Wei Ren Shaoheng Ni Kathrin M Bernt Jie Mi Nancy Kiviat André Lieber 《Molecular therapy》2004,9(4):496-509
We have constructed a new capsid-modified adenovirus (Ad) vector that specifically replicates in tumor cells and expresses TNF-related apoptosis-inducing ligand (TRAIL). The Ad capsid contains short-shafted fibers derived from Ad serotype 35, which allow for efficient infection of malignant tumor cells, and largely avoids innate toxicity after intravenous application. Replication-dependent homologous recombination in Ad genomes was used to achieve tumor-specific expression of Ad E1a (to mediate viral replication) and TRAIL (to mediate apoptosis and enhance release of progeny virus from infected cells). We demonstrated that our oncolytic vector (Ad5/35.IR-E1A/TRAIL) induced apoptosis in human tumor cell lines derived from colorectal, lung, prostate, and liver cancer. Both in vitro and in vivo tumor models showed efficient intratumoral spread of this vector. In a model for metastatic colon cancer, tail vein infusion of Ad5/35.IR-E1A/TRAIL resulted in elimination of preestablished liver metastases. Intravenous injection of this vector caused a transient elevation of serum glutamic pyruvic transaminase in tumor-bearing mice, which we attributed to factors released from apoptotic tumor cells. Liver histology analyzed at day 14 after virus injection did not show signs of hepatocellular damage. This new oncolytic vector represents a potentially efficient means for gene therapy of metastatic cancer. 相似文献
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Various neocortical areas from four females aged 16–24 years with Rett syndrome (RS) were investigated and compared with
brains of therapy-resistant partial epilepsy (TRPE) patients (18–25 years), infantile autism (IA), and control brains (24
and 58 years). The cytoarchitecture of area 10 (frontal), area 21 (temporal), area 4 (primary motor cortex), and area 17 (primary
visual cortex) was studied by the combined Klüver-Barrera (luxol fast blue and cresyl violet) standard procedure. Autofluorescence
of lipofuscin, immunofluorescence of synaptic vesicle proteins [synaptophysin (p38)] and lectin-stained (Wisteria floribunda agglutinin) perineuronal nets (PNs) were studied in the cortices using dual-channel confocal laser scanning microscopy. The
brains of RS females show various types of morphological/cytoarchitectonical abnormalities of single pyramidal neurons in
layers II–III, and V–VII of different cortical areas. The abnormalities include mild losses of pyramidal neurons, more pronounced
in layers II and III than in layers V and VII, and more evident in frontal and temporal areas than in the visual cortex. Microdysgenesis,
including abnormalities due to neuronal migration disorders, was not found in RS, in contrast to the observations in TRPE
patients, strongly indicating that RS is not a neuronal migration disorder. Lipofuscin distribution was normal but amounts
were lower in RS cases than in control and TRPE brains. PNs were less expressed in cortices of the IA case, but were clearly
overexpressed in the motor cortex of RS. Quantitative analysis of p38 showed a decrease in the area occupied by p38 immunoreactivity
by 20–40% in RS compared with controls. It is concluded that RS could best be explained by a postnatal synaptogenic developmental
deficiency; the basic defect, however, is still completely unknown.
Received: 26 February 1996 / Revised, accepted: 11 July 1996 相似文献
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Iwar Klime Antonian Vraana Jaroslav Kune Elena eboUkovaA Zdena Dobe ovaa Pavel tolba Josef Zicha 《Blood pressure》1995,4(3):137-142
Hereditary hypertriglyceridemic rats (hHTg) were developed as a new genetic model for the study of relationships between blood pressure (BP) and metabolic abnormalities. This strain has been produced by selective inbreeding from Wistar rats according to the rise of plasma triglycerides induced by a high-sucrose diet. Though hHTg rats display hypertriglyceridemia, impaired glucose tolerrance, hyperinsulinemia, insulin resistance and increased BP even without nutritional stimuli, high sucrose feeding further aggravates these symptoms. High plasma triglycerides levels in hHTg rats seem to be a consequence of their hyperproduction. Impaired insulin action is responsible for the defective glucoregulation in this strain. The loss of insulin responsiveness might be due to a reduction in the number of glucose transporters. Highly significant relationships among plasma triglycerides, ouabain-resistant Na+ transport and BP were demonstrated in the hHTg rats. Segregating populations (F2 hybrids) should be used for genetic analysis of the primary role of lipid and/or ion transport abnormalities in the pathogenesis of this form of genetic hypertension. 相似文献
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Oleg Kleiner Jacob Mordehai Pavel Krugliak Zahavi Cohen 《European Journal of Trauma》2004,30(6):403-405
Background:
Cases of extrahepatic biliary tree trauma are not as common as other intraabdominal injuries and may pose a diagnostic and therapeutic challenge. 相似文献10.
Pavel Rauch Pavel Rychetsky Igor Hochel Radovan Bilek Jean‐Luc Guesdon 《Food and Agricultural Immunology》1992,4(2):67-72
A competitive enzyme immunoassay of histamine in foodstuffs has been developed using a monoclonal antibody against a histamine‐benzoquinone adduct. In this assay, histamine present in food samples was treated with 1,4‐benzoquinone to form histamine‐benzoquinone by a simple and quick reaction and a histamine‐benzoquinone‐horse‐radish peroxidase conjugate was used as the labelled hapten. The apparent association constant (Ka) of the antibody used was 3.6 ×106 l/mol and Gibbs’ energy of the immune complex formation has been estimated to find the optimal incubation time of the assay. The method enabled determination of histamine in fish, cheese, wine and beer at a concentration as low as 7 ng/ml with an accuracy of ± 15%. The recovery of the immunoassay was 88.9–114%. Cross‐reactivities of histidine, tyramine, tryptamine and its derivatives were lower than 0.001% and did not affect the assay. 相似文献