Comparison of glycoproteins by two-dimensional mapping of glycosylated peptides

Salinovich, Olivia; Montelaro, Ronald C.

doi:10.1016/0003-2697(86)90190-9

Cited by 17 publications

(12 citation statements)

References 25 publications

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“…Equal amounts of proteins (25 or 50 µg) were separated by 10 or 12% SDS–polyacrylamide gel electrophoresis (PAGE) and transferred to nitrocellulose membranes. Equal lane loading was judged by non‐permanent Ponceau S staining of nitrocellulose membranes (Salinovich and Montelaro 1986). After rinsing to remove the staining, the membranes were blocked with 10% non‐fat milk and incubated with polyclonal anti‐iNOS (1 : 2500), anti‐COX‐2 (1 : 500) and anti‐mPGES (1 : 300) antibodies for 1 h at 25°C.…”

Section: Western Blot Analysismentioning

confidence: 99%

Prolonged exposure of microglia to lipopolysaccharide modifies the intracellular signaling pathways and selectively promotes prostaglandin E₂ synthesis

Ajmone‐Cat

Nicolini

Minghetti

2003

Journal of Neurochemistry

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During inflammatory or degenerative processes microglial cells are likely to be exposed to activating agents that persist in brain parenchyma for prolonged periods. As our knowledge on microglial activation is largely based on in vitro studies in which microglial cultures are activated by a single administration of pro-inflammatory stimuli, we investigated the effects of repeated endotoxin (LPS) challenges on microglial functional state. Primary rat microglial cultures were subjected to one, two or three consecutive LPS-stimulation and the production of tumor necrosis factor-a (TNF-a), nitric oxide (NO), prostaglandin E 2 (PGE 2 ) and 15-deoxy-D 12,14 -PGJ 2 (15d-PGJ 2 ) measured. The ability of microglial cells to produce NO, TNF-a and 15d-PGJ 2 upon the first LPS challenge rapidly declined after the second and the third stimulations, whereas PGE 2 synthesis remained constantly elevated. Accordingly, the expression of inducible NO synthase decreased whereas cyclooxygenase-2 and microsomal PGE synthase remained up-regulated. The signaling pathways evoked by single or multiple LPS-stimulation were also profoundly different, when considering the activation of the transcription factors nuclear factor-kappa B and CREB, and of the p38 MAPK. Our observations suggest that prolonged exposure to LPS, and likely other activating agents, induces in microglia a functional state clearly distinct from that triggered by acute stimulation. The progressive down-regulation of proinflammatory molecules and the sustained release of PGE 2 could have important implications for the resolution of brain inflammation.

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Section: Western Blot Analysismentioning

confidence: 99%

Prolonged exposure of microglia to lipopolysaccharide modifies the intracellular signaling pathways and selectively promotes prostaglandin E₂ synthesis

Ajmone‐Cat

Nicolini

Minghetti

2003

Journal of Neurochemistry

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“…Electroblotting was performed at 50 V overnight with stirring of the buffer [16]. Following transfer, the protein was visualized by staining the Hybond C filter with Ponceau S [0.2% (wh) Ponceau S; 7% acetic acid] for 5 min [17]. After destaining in water, to block nonspecific binding sites the filter was incubated in 5% FCS/PBS for 1 h at room temperature followed by a brief rinse in PBS.…”

Section: Western Blotsmentioning

confidence: 99%

The identification of the β₂‐microglobulin binding antigen encoded by the human CDID gene

Bilsland

Milstein

1991

Eur J Immunol

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Human cluster of differentiation (CD1) is a family of cell surface glycoproteins composed of a 43-49-kDa heavy chain non-covalently associated with beta 2-microglobulin. Five human CD1 genes have been detected and cloned. Three genes (CD1A, -B and -C) encode the serologically defined CD1a, -b and -c antigens. Thus two genes remain, CD1D and CD1E, whose protein products have not been characterized so far. This report describes how a beta-galactosidase-CD1D fusion protein was used to raise specific antisera and a monoclonal antibody against the CD1D gene product. The monoclonal antibody defines a cell surface molecule expressed on a cortical thymocyte cell line and is composed of a 49-kDa heavy chain associated with beta 2-microglobulin, which is serologically distinct from CD1a.

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“…The observation that SA had slightly higher nucleotide sequence identity (99.222%) with F2 than DE (99.158%) can probably be explained by the fact the latter foal survived almost three times as long post‐infection providing more time for nucleotide substitutions to accumulate. Examination of the distribution of the mutations present in viruses from the three foals demonstrated that overall, it closely resembled that seen in individual EIAV‐infected horses over time (Leroux, Issel, & Montelaro, ; Salinovich & Montelaro, ; Salinovich et al., ; Zheng et al., ) with most occurring in the env/ORF3 portion of the genome while gag, pol ORF, ORF2 and known transcription factor binding sites within the LTR possessed significantly higher levels of nucleotide sequence identity. Furthermore, the ratio of predicted synonymous to non‐synonymous nucleotide substitutions was approximately 2:1 in gag and pol compared with 1:20.5, 1:4.4 and 1:3.5 in sequences encoding SU, TM and the second exon of rev , respectively.…”

Section: Discussionmentioning

confidence: 82%

Deep sequencing and variant analysis of an Italian pathogenic field strain of equine infectious anaemia virus

Cappelli

Cook

Stefanetti

et al. 2017

Transbound Emerg Dis

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Equine infectious anaemia virus (EIAV) is a lentivirus with an almost worldwide distribution that causes persistent infections in equids. Technical limitations have restricted genetic analysis of EIAV field isolates predominantly to gag sequences resulting in very little published information concerning the extent of inter-strain variation in pol, env and the three ancillary open reading frames (ORFs). Here, we describe the use of long-range PCR in conjunction with next-generation sequencing (NGS) for rapid molecular characterization of all viral ORFs and known transcription factor binding motifs within the long terminal repeat of two EIAV isolates from the 2006 Italian outbreak. These isolates were from foals believed to have been exposed to the same source material but with different clinical histories: one died 53 days post-infection (SA) while the other (DE) survived 5 months despite experiencing multiple febrile episodes. Nucleotide sequence identity between the isolates was 99.358% confirming infection with the same EIAV strain with most differences comprising single nucleotide polymorphisms in env and the second exon of rev. Although the synonymous:non-synonymous nucleotide substitution ratio was approximately 2:1 in gag and pol, the situation is reversed in env and ORF3 suggesting these sequences are subjected to host-mediated selective pressure. EIAV proviral quasispecies complexity in vivo has not been extensively investigated; however, analysis suggests it was relatively low in SA at the time of death. These results highlight advantages of NGS for molecular characterization of EIAV namely it avoids potential artefacts generated by traditional composite sequencing strategies and can provide information about viral quasispecies complexity.

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Comparison of glycoproteins by two-dimensional mapping of glycosylated peptides

Cited by 17 publications

References 25 publications

Prolonged exposure of microglia to lipopolysaccharide modifies the intracellular signaling pathways and selectively promotes prostaglandin E₂ synthesis

Prolonged exposure of microglia to lipopolysaccharide modifies the intracellular signaling pathways and selectively promotes prostaglandin E₂ synthesis

The identification of the β₂‐microglobulin binding antigen encoded by the human CDID gene

Deep sequencing and variant analysis of an Italian pathogenic field strain of equine infectious anaemia virus

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Comparison of glycoproteins by two-dimensional mapping of glycosylated peptides

Cited by 17 publications

References 25 publications

Prolonged exposure of microglia to lipopolysaccharide modifies the intracellular signaling pathways and selectively promotes prostaglandin E2 synthesis

Prolonged exposure of microglia to lipopolysaccharide modifies the intracellular signaling pathways and selectively promotes prostaglandin E2 synthesis

The identification of the β2‐microglobulin binding antigen encoded by the human CDID gene

Deep sequencing and variant analysis of an Italian pathogenic field strain of equine infectious anaemia virus

Contact Info

Product

Resources

About

Prolonged exposure of microglia to lipopolysaccharide modifies the intracellular signaling pathways and selectively promotes prostaglandin E₂ synthesis

Prolonged exposure of microglia to lipopolysaccharide modifies the intracellular signaling pathways and selectively promotes prostaglandin E₂ synthesis

The identification of the β₂‐microglobulin binding antigen encoded by the human CDID gene