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Roche nonidet p40 lysis buffer
Nonidet P40 Lysis Buffer, supplied by Roche, used in various techniques. Bioz Stars score: 93/100, based on 12 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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nonidet p40 lysis buffer - by Bioz Stars, 2020-04
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Centrifugation:

Article Title: Endogenous RGS14 is a cytoplasmic-nuclear shuttling protein that localizes to juxtanuclear membranes and chromatin-rich regions of the nucleus
Article Snippet: Cells were then resuspended in 10 volumes of Nonidet-P40 lysis buffer (10 mM HEPES, pH 7.5; 10 mM KCl; 0.1 mM EDTA; 1 mM dithiothreitol (DTT); 0.5% Nonidet‐40; protease inhibitor cocktail (Roche)) and allowed to swell in ice for 12 min with intermittent mixing. .. After centrifugation at 1,200 g for 8 min, the supernatant was collected as cytoplasmic extract and supplemented with 240 mM NaCl.

Article Title: Erythroid Progenitor Cells in Atlantic Salmon (Salmo salar) May Be Persistently and Productively Infected with Piscine Orthoreovirus (PRV)
Article Snippet: For each sample, 15 µL blood pellet was added to Nonidet-P40 lysis buffer containing complete ultra mini protease inhibitor cocktail (1:5) (Roche, Mannheim, Germany) and incubated for 30 min on ice. .. After centrifugation at 5000× g for 5 min, the supernatant was diluted with DEPC treated water (1:5) and mixed with XT sample buffer and XT reducing agent (Bio-Rad, Hercules, CA, USA) and boiled at 95 °C for 5 min. PRV proteins were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), using 4–12% criterion XT bis-tris gel.

Article Title: The non-structural protein μNS of piscine orthoreovirus (PRV) forms viral factory-like structures
Article Snippet: Immunoprecipitation A total of 5 million EPC cells were pelleted by centrifugation, resuspended in 100 μL Ingenio Electroporation Solution (Mirus, Madison, WI, USA) and co-transfected with 8 μg plasmid using the Amaxa T-20 program. pcDNA3.1-μNS-N-FLAG was co-transfected with pcDNA3.1-σNS-N-MYC, pcDNA3.1-μ2-C-HA, pcDNA3.1-λ1-N-HA and pcDNA3.1 S8ORF2 (negative control) separately, using three parallel preparations. .. Cells were collected from the culture flasks 72 h post transfection (hpt), centrifuged at 5000 g for 5 min and resuspended in 1 mL Nonidet-P40 lysis buffer (1% NP-40, 50 mM Tris–HCl pH 8.0, 150 mM NaCl, 2 mM EDTA) containing Complete ultra mini protease inhibitor cocktail (Roche, Mannheim, Germany).

Article Title: Viral Protein Kinetics of Piscine Orthoreovirus Infection in Atlantic Salmon Blood Cells
Article Snippet: A total of 3 × 5 million EPC cells were pelleted by centrifugation, resuspended in 100 μL Ingenio Electroporation Solution (Mirus, Madison, WI, USA) and transfected with 4 μg pcDNA3.1 µNS N-FLAG or pcDNA3.1 λ1 N-HA. .. The cell pellets were lysed in Nonidet-P40 lysis buffer (1% NP-40, 50 mM Tris–HCl pH 8.0, 150 mM NaCl, 2 mM EDTA) containing Complete ultra mini protease inhibitor cocktail (Roche, Mannheim, Germany).

Electrophoresis:

Article Title: LPS-induced IL-8 activation in human intestinal epithelial cells is accompanied by specific histone H3 acetylation and methylation changes
Article Snippet: Western Blot Analysis Cell extracts were prepared in Nonidet P40 lysis buffer with 1 mM PMSF and Complete™ protease inhibitors mix (Roche, Indianapolis, IN, USA). .. 50 μg of proteins were resolved by electrophoresis using 10% SDS-PAGE gels and transferred to BA 85 0.45 μm PROTAN nitrocellulose filters (Schleicher & Schnell, Inc., Dassel, Germany).

Incubation:

Article Title: Erythroid Progenitor Cells in Atlantic Salmon (Salmo salar) May Be Persistently and Productively Infected with Piscine Orthoreovirus (PRV)
Article Snippet: .. For each sample, 15 µL blood pellet was added to Nonidet-P40 lysis buffer containing complete ultra mini protease inhibitor cocktail (1:5) (Roche, Mannheim, Germany) and incubated for 30 min on ice. .. After centrifugation at 5000× g for 5 min, the supernatant was diluted with DEPC treated water (1:5) and mixed with XT sample buffer and XT reducing agent (Bio-Rad, Hercules, CA, USA) and boiled at 95 °C for 5 min. PRV proteins were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), using 4–12% criterion XT bis-tris gel.

Article Title: LPS-induced IL-8 activation in human intestinal epithelial cells is accompanied by specific histone H3 acetylation and methylation changes
Article Snippet: Western Blot Analysis Cell extracts were prepared in Nonidet P40 lysis buffer with 1 mM PMSF and Complete™ protease inhibitors mix (Roche, Indianapolis, IN, USA). .. The blots were incubated with rabbit anti-IκB-α antibodies (Santa Cruz Biotechnology, Santa Cruz, CA, USA) and mouse anti-γ-tubulin antibodies (Sigma-Aldrich Corp. St. Louis, MO, USA) as a control for protein loading.

Article Title: LZAP Inhibits p38 MAPK (p38) Phosphorylation and Activity by Facilitating p38 Association with the Wild-Type p53 Induced Phosphatase 1 (WIP1)
Article Snippet: Immunoprecipitation and immunoblotting Cells were lysed in 0.5%(v/v) Nonidet P40 lysis buffer supplemented with protease inhibitor cocktail (Roche). .. Total cell extracts were incubated with specific antibodies and precipitated with protein A or G sepharose beads (GE Healthcare) before washing and suspension in Laemmli and gel electrophoresis followed by immunoblotting as described

Article Title: Molecular interactions of Bcl-2 and Bcl-xL with mortalin: identification and functional characterization
Article Snippet: Cells (U2OS; osteosarcoma) were cultured in DMEM (Dulbecco's modified Eagle's medium) essential medium supplemented with 10% (v/v) FBS (fetal bovine serum) and incubated in a 5% CO2 /95% air humidified incubator. .. Cell lysates were prepared in Nonidet P40 lysis buffer [10 mM Tris/HCl (pH 7.4), 150 mM NaCl, 5 mM EDTA, 1% Nonidet P40] supplemented with a protease inhibitor cocktail (Complete™ Mini; Roche Diagnostics) following 48 h of transfection.

Article Title: The non-structural protein μNS of piscine orthoreovirus (PRV) forms viral factory-like structures
Article Snippet: Cells were collected from the culture flasks 72 h post transfection (hpt), centrifuged at 5000 g for 5 min and resuspended in 1 mL Nonidet-P40 lysis buffer (1% NP-40, 50 mM Tris–HCl pH 8.0, 150 mM NaCl, 2 mM EDTA) containing Complete ultra mini protease inhibitor cocktail (Roche, Mannheim, Germany). .. The supernatant was transferred to a new tube, added antibodies against the desired epitope tag or anti-S8ORF2 and incubated overnight at 4 °C with rotation.

Article Title: Wip1 phosphatase modulates both long-term potentiation and long-term depression through the dephosphorylation of CaMKII
Article Snippet: Tissues were lysed in 0.5% (v/v) Nonidet P40 lysis buffer supplemented with a protease inhibitor cocktail and PhosStop (Roche). .. The total cell extracts were incubated with specific antibodies and precipitated with protein A/G sepharose beads (sigma) before washing and suspension in Laemmli.

Infection:

Article Title: Erythroid Progenitor Cells in Atlantic Salmon (Salmo salar) May Be Persistently and Productively Infected with Piscine Orthoreovirus (PRV)
Article Snippet: Heparinized blood from PRV-1 infected and uninfected fish from a previous challenge trial were used as positive and negative controls, respectively [ ]. .. For each sample, 15 µL blood pellet was added to Nonidet-P40 lysis buffer containing complete ultra mini protease inhibitor cocktail (1:5) (Roche, Mannheim, Germany) and incubated for 30 min on ice.

Expressing:

Article Title: Erythroid Progenitor Cells in Atlantic Salmon (Salmo salar) May Be Persistently and Productively Infected with Piscine Orthoreovirus (PRV)
Article Snippet: Western Blotting Heparinized blood cell pellets from the experimental PRV-1 challenged Atlantic salmon were used for virus protein expression analysis in western blotting (WB). .. For each sample, 15 µL blood pellet was added to Nonidet-P40 lysis buffer containing complete ultra mini protease inhibitor cocktail (1:5) (Roche, Mannheim, Germany) and incubated for 30 min on ice.

Article Title: Molecular interactions of Bcl-2 and Bcl-xL with mortalin: identification and functional characterization
Article Snippet: Protein expression was observed as either direct GFP fluorescence under the microscope or by Western blotting with anti-GFP antibody. .. Cell lysates were prepared in Nonidet P40 lysis buffer [10 mM Tris/HCl (pH 7.4), 150 mM NaCl, 5 mM EDTA, 1% Nonidet P40] supplemented with a protease inhibitor cocktail (Complete™ Mini; Roche Diagnostics) following 48 h of transfection.

Article Title: The non-structural protein μNS of piscine orthoreovirus (PRV) forms viral factory-like structures
Article Snippet: From each culture flask, 0.5 mL transfected cells were transferred to a 24-well plate intended for expression analysis by immunofluorescence microscopy. .. Cells were collected from the culture flasks 72 h post transfection (hpt), centrifuged at 5000 g for 5 min and resuspended in 1 mL Nonidet-P40 lysis buffer (1% NP-40, 50 mM Tris–HCl pH 8.0, 150 mM NaCl, 2 mM EDTA) containing Complete ultra mini protease inhibitor cocktail (Roche, Mannheim, Germany).

Modification:

Article Title: Endogenous RGS14 is a cytoplasmic-nuclear shuttling protein that localizes to juxtanuclear membranes and chromatin-rich regions of the nucleus
Article Snippet: Subcellular fractionation B35 cells were lysed and fractioned to isolate intact nuclei and cytosol in a protocol modified from [ ]. .. Cells were then resuspended in 10 volumes of Nonidet-P40 lysis buffer (10 mM HEPES, pH 7.5; 10 mM KCl; 0.1 mM EDTA; 1 mM dithiothreitol (DTT); 0.5% Nonidet‐40; protease inhibitor cocktail (Roche)) and allowed to swell in ice for 12 min with intermittent mixing.

Article Title: Molecular interactions of Bcl-2 and Bcl-xL with mortalin: identification and functional characterization
Article Snippet: Cells (U2OS; osteosarcoma) were cultured in DMEM (Dulbecco's modified Eagle's medium) essential medium supplemented with 10% (v/v) FBS (fetal bovine serum) and incubated in a 5% CO2 /95% air humidified incubator. .. Cell lysates were prepared in Nonidet P40 lysis buffer [10 mM Tris/HCl (pH 7.4), 150 mM NaCl, 5 mM EDTA, 1% Nonidet P40] supplemented with a protease inhibitor cocktail (Complete™ Mini; Roche Diagnostics) following 48 h of transfection.

Western Blot:

Article Title: Erythroid Progenitor Cells in Atlantic Salmon (Salmo salar) May Be Persistently and Productively Infected with Piscine Orthoreovirus (PRV)
Article Snippet: Paragraph title: 2.6. Western Blotting ... For each sample, 15 µL blood pellet was added to Nonidet-P40 lysis buffer containing complete ultra mini protease inhibitor cocktail (1:5) (Roche, Mannheim, Germany) and incubated for 30 min on ice.

Article Title: LPS-induced IL-8 activation in human intestinal epithelial cells is accompanied by specific histone H3 acetylation and methylation changes
Article Snippet: .. Western Blot Analysis Cell extracts were prepared in Nonidet P40 lysis buffer with 1 mM PMSF and Complete™ protease inhibitors mix (Roche, Indianapolis, IN, USA). .. 50 μg of proteins were resolved by electrophoresis using 10% SDS-PAGE gels and transferred to BA 85 0.45 μm PROTAN nitrocellulose filters (Schleicher & Schnell, Inc., Dassel, Germany).

Article Title: Caspase-mediated cleavage of the exosome subunit PM/Scl-75 during apoptosis
Article Snippet: .. Western blot analysis Cells were lysed on ice for 30 minutes in Nonidet P40 lysis buffer (25 mM Tris-HCl, pH 7.5, 1% Nonidet P40, 100 mM KCl, 10 mM MgCl2 , 1 mM dithiothreitol), containing a protease inhibitor cocktail (Complete; Roche, Mannheim, Germany). ..

Article Title: Molecular interactions of Bcl-2 and Bcl-xL with mortalin: identification and functional characterization
Article Snippet: Protein expression was observed as either direct GFP fluorescence under the microscope or by Western blotting with anti-GFP antibody. .. Cell lysates were prepared in Nonidet P40 lysis buffer [10 mM Tris/HCl (pH 7.4), 150 mM NaCl, 5 mM EDTA, 1% Nonidet P40] supplemented with a protease inhibitor cocktail (Complete™ Mini; Roche Diagnostics) following 48 h of transfection.

Article Title: Viral Protein Kinetics of Piscine Orthoreovirus Infection in Atlantic Salmon Blood Cells
Article Snippet: Transfected EPC cells were also used to further verify anti-µNS and anti-λ1 in WB. .. The cell pellets were lysed in Nonidet-P40 lysis buffer (1% NP-40, 50 mM Tris–HCl pH 8.0, 150 mM NaCl, 2 mM EDTA) containing Complete ultra mini protease inhibitor cocktail (Roche, Mannheim, Germany).

Article Title: Wip1 phosphatase modulates both long-term potentiation and long-term depression through the dephosphorylation of CaMKII
Article Snippet: Tissues were lysed in 0.5% (v/v) Nonidet P40 lysis buffer supplemented with a protease inhibitor cocktail and PhosStop (Roche). .. Tissues were lysed in 0.5% (v/v) Nonidet P40 lysis buffer supplemented with a protease inhibitor cocktail and PhosStop (Roche).

Electroporation:

Article Title: The non-structural protein μNS of piscine orthoreovirus (PRV) forms viral factory-like structures
Article Snippet: Immunoprecipitation A total of 5 million EPC cells were pelleted by centrifugation, resuspended in 100 μL Ingenio Electroporation Solution (Mirus, Madison, WI, USA) and co-transfected with 8 μg plasmid using the Amaxa T-20 program. pcDNA3.1-μNS-N-FLAG was co-transfected with pcDNA3.1-σNS-N-MYC, pcDNA3.1-μ2-C-HA, pcDNA3.1-λ1-N-HA and pcDNA3.1 S8ORF2 (negative control) separately, using three parallel preparations. .. Cells were collected from the culture flasks 72 h post transfection (hpt), centrifuged at 5000 g for 5 min and resuspended in 1 mL Nonidet-P40 lysis buffer (1% NP-40, 50 mM Tris–HCl pH 8.0, 150 mM NaCl, 2 mM EDTA) containing Complete ultra mini protease inhibitor cocktail (Roche, Mannheim, Germany).

Article Title: Viral Protein Kinetics of Piscine Orthoreovirus Infection in Atlantic Salmon Blood Cells
Article Snippet: A total of 3 × 5 million EPC cells were pelleted by centrifugation, resuspended in 100 μL Ingenio Electroporation Solution (Mirus, Madison, WI, USA) and transfected with 4 μg pcDNA3.1 µNS N-FLAG or pcDNA3.1 λ1 N-HA. .. The cell pellets were lysed in Nonidet-P40 lysis buffer (1% NP-40, 50 mM Tris–HCl pH 8.0, 150 mM NaCl, 2 mM EDTA) containing Complete ultra mini protease inhibitor cocktail (Roche, Mannheim, Germany).

Transfection:

Article Title: WDRPUH, A Novel WD-Repeat-Containing Protein, Is Highly Expressed in Human Hepatocellular Carcinoma and Involved in Cell Proliferation 1WDRPUH, A Novel WD-Repeat-Containing Protein, Is Highly Expressed in Human Hepatocellular Carcinoma and Involved in Cell Proliferation 1 *
Article Snippet: .. The cells were harvested at 48 hours after transfection and lysed with Nonidet P40 lysis buffer [20 mM Tris-HCl pH 7.5, 150 mM NaCl, 1% NP-40, 1x complete Protease Inhibitor Cocktail EDTA (-) (Roche, Basel, Switzerland)]. .. Cell lysates were immunoprecipitated with anti-FLAG M2 antibody conjugated with agarose overnight at 4°C.

Article Title: Molecular interactions of Bcl-2 and Bcl-xL with mortalin: identification and functional characterization
Article Snippet: .. Cell lysates were prepared in Nonidet P40 lysis buffer [10 mM Tris/HCl (pH 7.4), 150 mM NaCl, 5 mM EDTA, 1% Nonidet P40] supplemented with a protease inhibitor cocktail (Complete™ Mini; Roche Diagnostics) following 48 h of transfection. .. Stable cells lines were obtained by 2 weeks of selection for the transfected gene using 800 μg/ml Gly418 (Invitrogen) in the medium, and then maintained with 200 μg/ml Gly418 in DMEM.

Article Title: The non-structural protein μNS of piscine orthoreovirus (PRV) forms viral factory-like structures
Article Snippet: .. Cells were collected from the culture flasks 72 h post transfection (hpt), centrifuged at 5000 g for 5 min and resuspended in 1 mL Nonidet-P40 lysis buffer (1% NP-40, 50 mM Tris–HCl pH 8.0, 150 mM NaCl, 2 mM EDTA) containing Complete ultra mini protease inhibitor cocktail (Roche, Mannheim, Germany). .. The supernatant was transferred to a new tube, added antibodies against the desired epitope tag or anti-S8ORF2 and incubated overnight at 4 °C with rotation.

Article Title: Viral Protein Kinetics of Piscine Orthoreovirus Infection in Atlantic Salmon Blood Cells
Article Snippet: The transfected cells were transferred to 75 cm2 culture flasks containing 20 mL pre-equilibrated L-15 growth medium (described above) and collected 72 h post-transfection. .. The cell pellets were lysed in Nonidet-P40 lysis buffer (1% NP-40, 50 mM Tris–HCl pH 8.0, 150 mM NaCl, 2 mM EDTA) containing Complete ultra mini protease inhibitor cocktail (Roche, Mannheim, Germany).

Protease Inhibitor:

Article Title: Endogenous RGS14 is a cytoplasmic-nuclear shuttling protein that localizes to juxtanuclear membranes and chromatin-rich regions of the nucleus
Article Snippet: .. Cells were then resuspended in 10 volumes of Nonidet-P40 lysis buffer (10 mM HEPES, pH 7.5; 10 mM KCl; 0.1 mM EDTA; 1 mM dithiothreitol (DTT); 0.5% Nonidet‐40; protease inhibitor cocktail (Roche)) and allowed to swell in ice for 12 min with intermittent mixing. .. Samples were then vortexed at max speed for 10–12 sec to disrupt cell membranes, and 10% of the volume was removed for later assessing whole cell lysates by immunoblotting.

Article Title: WDRPUH, A Novel WD-Repeat-Containing Protein, Is Highly Expressed in Human Hepatocellular Carcinoma and Involved in Cell Proliferation 1WDRPUH, A Novel WD-Repeat-Containing Protein, Is Highly Expressed in Human Hepatocellular Carcinoma and Involved in Cell Proliferation 1 *
Article Snippet: .. The cells were harvested at 48 hours after transfection and lysed with Nonidet P40 lysis buffer [20 mM Tris-HCl pH 7.5, 150 mM NaCl, 1% NP-40, 1x complete Protease Inhibitor Cocktail EDTA (-) (Roche, Basel, Switzerland)]. .. Cell lysates were immunoprecipitated with anti-FLAG M2 antibody conjugated with agarose overnight at 4°C.

Article Title: Erythroid Progenitor Cells in Atlantic Salmon (Salmo salar) May Be Persistently and Productively Infected with Piscine Orthoreovirus (PRV)
Article Snippet: .. For each sample, 15 µL blood pellet was added to Nonidet-P40 lysis buffer containing complete ultra mini protease inhibitor cocktail (1:5) (Roche, Mannheim, Germany) and incubated for 30 min on ice. .. After centrifugation at 5000× g for 5 min, the supernatant was diluted with DEPC treated water (1:5) and mixed with XT sample buffer and XT reducing agent (Bio-Rad, Hercules, CA, USA) and boiled at 95 °C for 5 min. PRV proteins were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), using 4–12% criterion XT bis-tris gel.

Article Title: Caspase-mediated cleavage of the exosome subunit PM/Scl-75 during apoptosis
Article Snippet: .. Western blot analysis Cells were lysed on ice for 30 minutes in Nonidet P40 lysis buffer (25 mM Tris-HCl, pH 7.5, 1% Nonidet P40, 100 mM KCl, 10 mM MgCl2 , 1 mM dithiothreitol), containing a protease inhibitor cocktail (Complete; Roche, Mannheim, Germany). ..

Article Title: LZAP Inhibits p38 MAPK (p38) Phosphorylation and Activity by Facilitating p38 Association with the Wild-Type p53 Induced Phosphatase 1 (WIP1)
Article Snippet: .. Immunoprecipitation and immunoblotting Cells were lysed in 0.5%(v/v) Nonidet P40 lysis buffer supplemented with protease inhibitor cocktail (Roche). .. Total cell extracts were incubated with specific antibodies and precipitated with protein A or G sepharose beads (GE Healthcare) before washing and suspension in Laemmli and gel electrophoresis followed by immunoblotting as described

Article Title: Molecular interactions of Bcl-2 and Bcl-xL with mortalin: identification and functional characterization
Article Snippet: .. Cell lysates were prepared in Nonidet P40 lysis buffer [10 mM Tris/HCl (pH 7.4), 150 mM NaCl, 5 mM EDTA, 1% Nonidet P40] supplemented with a protease inhibitor cocktail (Complete™ Mini; Roche Diagnostics) following 48 h of transfection. .. Stable cells lines were obtained by 2 weeks of selection for the transfected gene using 800 μg/ml Gly418 (Invitrogen) in the medium, and then maintained with 200 μg/ml Gly418 in DMEM.

Article Title: The non-structural protein μNS of piscine orthoreovirus (PRV) forms viral factory-like structures
Article Snippet: .. Cells were collected from the culture flasks 72 h post transfection (hpt), centrifuged at 5000 g for 5 min and resuspended in 1 mL Nonidet-P40 lysis buffer (1% NP-40, 50 mM Tris–HCl pH 8.0, 150 mM NaCl, 2 mM EDTA) containing Complete ultra mini protease inhibitor cocktail (Roche, Mannheim, Germany). .. The supernatant was transferred to a new tube, added antibodies against the desired epitope tag or anti-S8ORF2 and incubated overnight at 4 °C with rotation.

Article Title: Viral Protein Kinetics of Piscine Orthoreovirus Infection in Atlantic Salmon Blood Cells
Article Snippet: .. The cell pellets were lysed in Nonidet-P40 lysis buffer (1% NP-40, 50 mM Tris–HCl pH 8.0, 150 mM NaCl, 2 mM EDTA) containing Complete ultra mini protease inhibitor cocktail (Roche, Mannheim, Germany). .. The supernatant was mixed with Sample Buffer (Bio-Rad) and Reducing Agent (Bio-Rad), denatured for 5 min at 95 °C and run in SDS-PAGE, using 4%–12% Bis–Tris Criterion XT gel (Bio-Rad).

Article Title: Wip1 phosphatase modulates both long-term potentiation and long-term depression through the dephosphorylation of CaMKII
Article Snippet: .. Tissues were lysed in 0.5% (v/v) Nonidet P40 lysis buffer supplemented with a protease inhibitor cocktail and PhosStop (Roche). .. The total cell extracts were incubated with specific antibodies and precipitated with protein A/G sepharose beads (sigma) before washing and suspension in Laemmli.

Cell Culture:

Article Title: Molecular interactions of Bcl-2 and Bcl-xL with mortalin: identification and functional characterization
Article Snippet: Paragraph title: Plasmids, cell culture, transfections and antibodies ... Cell lysates were prepared in Nonidet P40 lysis buffer [10 mM Tris/HCl (pH 7.4), 150 mM NaCl, 5 mM EDTA, 1% Nonidet P40] supplemented with a protease inhibitor cocktail (Complete™ Mini; Roche Diagnostics) following 48 h of transfection.

Immunofluorescence:

Article Title: The non-structural protein μNS of piscine orthoreovirus (PRV) forms viral factory-like structures
Article Snippet: From each culture flask, 0.5 mL transfected cells were transferred to a 24-well plate intended for expression analysis by immunofluorescence microscopy. .. Cells were collected from the culture flasks 72 h post transfection (hpt), centrifuged at 5000 g for 5 min and resuspended in 1 mL Nonidet-P40 lysis buffer (1% NP-40, 50 mM Tris–HCl pH 8.0, 150 mM NaCl, 2 mM EDTA) containing Complete ultra mini protease inhibitor cocktail (Roche, Mannheim, Germany).

Nucleic Acid Electrophoresis:

Article Title: Erythroid Progenitor Cells in Atlantic Salmon (Salmo salar) May Be Persistently and Productively Infected with Piscine Orthoreovirus (PRV)
Article Snippet: For each sample, 15 µL blood pellet was added to Nonidet-P40 lysis buffer containing complete ultra mini protease inhibitor cocktail (1:5) (Roche, Mannheim, Germany) and incubated for 30 min on ice. .. After centrifugation at 5000× g for 5 min, the supernatant was diluted with DEPC treated water (1:5) and mixed with XT sample buffer and XT reducing agent (Bio-Rad, Hercules, CA, USA) and boiled at 95 °C for 5 min. PRV proteins were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), using 4–12% criterion XT bis-tris gel.

Article Title: LZAP Inhibits p38 MAPK (p38) Phosphorylation and Activity by Facilitating p38 Association with the Wild-Type p53 Induced Phosphatase 1 (WIP1)
Article Snippet: Immunoprecipitation and immunoblotting Cells were lysed in 0.5%(v/v) Nonidet P40 lysis buffer supplemented with protease inhibitor cocktail (Roche). .. Total cell extracts were incubated with specific antibodies and precipitated with protein A or G sepharose beads (GE Healthcare) before washing and suspension in Laemmli and gel electrophoresis followed by immunoblotting as described

Fluorescence:

Article Title: Molecular interactions of Bcl-2 and Bcl-xL with mortalin: identification and functional characterization
Article Snippet: Protein expression was observed as either direct GFP fluorescence under the microscope or by Western blotting with anti-GFP antibody. .. Cell lysates were prepared in Nonidet P40 lysis buffer [10 mM Tris/HCl (pH 7.4), 150 mM NaCl, 5 mM EDTA, 1% Nonidet P40] supplemented with a protease inhibitor cocktail (Complete™ Mini; Roche Diagnostics) following 48 h of transfection.

Article Title: Viral Protein Kinetics of Piscine Orthoreovirus Infection in Atlantic Salmon Blood Cells
Article Snippet: The cell pellets were lysed in Nonidet-P40 lysis buffer (1% NP-40, 50 mM Tris–HCl pH 8.0, 150 mM NaCl, 2 mM EDTA) containing Complete ultra mini protease inhibitor cocktail (Roche, Mannheim, Germany). .. The cell pellets were lysed in Nonidet-P40 lysis buffer (1% NP-40, 50 mM Tris–HCl pH 8.0, 150 mM NaCl, 2 mM EDTA) containing Complete ultra mini protease inhibitor cocktail (Roche, Mannheim, Germany).

Size-exclusion Chromatography:

Article Title: Endogenous RGS14 is a cytoplasmic-nuclear shuttling protein that localizes to juxtanuclear membranes and chromatin-rich regions of the nucleus
Article Snippet: Cells were then resuspended in 10 volumes of Nonidet-P40 lysis buffer (10 mM HEPES, pH 7.5; 10 mM KCl; 0.1 mM EDTA; 1 mM dithiothreitol (DTT); 0.5% Nonidet‐40; protease inhibitor cocktail (Roche)) and allowed to swell in ice for 12 min with intermittent mixing. .. Samples were then vortexed at max speed for 10–12 sec to disrupt cell membranes, and 10% of the volume was removed for later assessing whole cell lysates by immunoblotting.

Microscopy:

Article Title: Molecular interactions of Bcl-2 and Bcl-xL with mortalin: identification and functional characterization
Article Snippet: Protein expression was observed as either direct GFP fluorescence under the microscope or by Western blotting with anti-GFP antibody. .. Cell lysates were prepared in Nonidet P40 lysis buffer [10 mM Tris/HCl (pH 7.4), 150 mM NaCl, 5 mM EDTA, 1% Nonidet P40] supplemented with a protease inhibitor cocktail (Complete™ Mini; Roche Diagnostics) following 48 h of transfection.

Article Title: The non-structural protein μNS of piscine orthoreovirus (PRV) forms viral factory-like structures
Article Snippet: From each culture flask, 0.5 mL transfected cells were transferred to a 24-well plate intended for expression analysis by immunofluorescence microscopy. .. Cells were collected from the culture flasks 72 h post transfection (hpt), centrifuged at 5000 g for 5 min and resuspended in 1 mL Nonidet-P40 lysis buffer (1% NP-40, 50 mM Tris–HCl pH 8.0, 150 mM NaCl, 2 mM EDTA) containing Complete ultra mini protease inhibitor cocktail (Roche, Mannheim, Germany).

Article Title: Viral Protein Kinetics of Piscine Orthoreovirus Infection in Atlantic Salmon Blood Cells
Article Snippet: The cell pellets were lysed in Nonidet-P40 lysis buffer (1% NP-40, 50 mM Tris–HCl pH 8.0, 150 mM NaCl, 2 mM EDTA) containing Complete ultra mini protease inhibitor cocktail (Roche, Mannheim, Germany). .. The cell pellets were lysed in Nonidet-P40 lysis buffer (1% NP-40, 50 mM Tris–HCl pH 8.0, 150 mM NaCl, 2 mM EDTA) containing Complete ultra mini protease inhibitor cocktail (Roche, Mannheim, Germany).

Protein Extraction:

Article Title: The non-structural protein μNS of piscine orthoreovirus (PRV) forms viral factory-like structures
Article Snippet: Cells were collected from the culture flasks 72 h post transfection (hpt), centrifuged at 5000 g for 5 min and resuspended in 1 mL Nonidet-P40 lysis buffer (1% NP-40, 50 mM Tris–HCl pH 8.0, 150 mM NaCl, 2 mM EDTA) containing Complete ultra mini protease inhibitor cocktail (Roche, Mannheim, Germany). .. The Immunoprecipitation Kit Dynabeads Protein G (Novex, Life Technologies) was used for protein extraction and the beads prepared according to the manufacturer’s protocol.

Polyacrylamide Gel Electrophoresis:

Article Title: WDRPUH, A Novel WD-Repeat-Containing Protein, Is Highly Expressed in Human Hepatocellular Carcinoma and Involved in Cell Proliferation 1WDRPUH, A Novel WD-Repeat-Containing Protein, Is Highly Expressed in Human Hepatocellular Carcinoma and Involved in Cell Proliferation 1 *
Article Snippet: The cells were harvested at 48 hours after transfection and lysed with Nonidet P40 lysis buffer [20 mM Tris-HCl pH 7.5, 150 mM NaCl, 1% NP-40, 1x complete Protease Inhibitor Cocktail EDTA (-) (Roche, Basel, Switzerland)]. .. The eluted complex was centrifuged and supernatant was resuspended in 5x Laemmli sample buffer and separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE).

Lysis:

Article Title: Endogenous RGS14 is a cytoplasmic-nuclear shuttling protein that localizes to juxtanuclear membranes and chromatin-rich regions of the nucleus
Article Snippet: .. Cells were then resuspended in 10 volumes of Nonidet-P40 lysis buffer (10 mM HEPES, pH 7.5; 10 mM KCl; 0.1 mM EDTA; 1 mM dithiothreitol (DTT); 0.5% Nonidet‐40; protease inhibitor cocktail (Roche)) and allowed to swell in ice for 12 min with intermittent mixing. .. Samples were then vortexed at max speed for 10–12 sec to disrupt cell membranes, and 10% of the volume was removed for later assessing whole cell lysates by immunoblotting.

Article Title: WDRPUH, A Novel WD-Repeat-Containing Protein, Is Highly Expressed in Human Hepatocellular Carcinoma and Involved in Cell Proliferation 1WDRPUH, A Novel WD-Repeat-Containing Protein, Is Highly Expressed in Human Hepatocellular Carcinoma and Involved in Cell Proliferation 1 *
Article Snippet: .. The cells were harvested at 48 hours after transfection and lysed with Nonidet P40 lysis buffer [20 mM Tris-HCl pH 7.5, 150 mM NaCl, 1% NP-40, 1x complete Protease Inhibitor Cocktail EDTA (-) (Roche, Basel, Switzerland)]. .. Cell lysates were immunoprecipitated with anti-FLAG M2 antibody conjugated with agarose overnight at 4°C.

Article Title: Erythroid Progenitor Cells in Atlantic Salmon (Salmo salar) May Be Persistently and Productively Infected with Piscine Orthoreovirus (PRV)
Article Snippet: .. For each sample, 15 µL blood pellet was added to Nonidet-P40 lysis buffer containing complete ultra mini protease inhibitor cocktail (1:5) (Roche, Mannheim, Germany) and incubated for 30 min on ice. .. After centrifugation at 5000× g for 5 min, the supernatant was diluted with DEPC treated water (1:5) and mixed with XT sample buffer and XT reducing agent (Bio-Rad, Hercules, CA, USA) and boiled at 95 °C for 5 min. PRV proteins were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), using 4–12% criterion XT bis-tris gel.

Article Title: LPS-induced IL-8 activation in human intestinal epithelial cells is accompanied by specific histone H3 acetylation and methylation changes
Article Snippet: .. Western Blot Analysis Cell extracts were prepared in Nonidet P40 lysis buffer with 1 mM PMSF and Complete™ protease inhibitors mix (Roche, Indianapolis, IN, USA). .. 50 μg of proteins were resolved by electrophoresis using 10% SDS-PAGE gels and transferred to BA 85 0.45 μm PROTAN nitrocellulose filters (Schleicher & Schnell, Inc., Dassel, Germany).

Article Title: Caspase-mediated cleavage of the exosome subunit PM/Scl-75 during apoptosis
Article Snippet: .. Western blot analysis Cells were lysed on ice for 30 minutes in Nonidet P40 lysis buffer (25 mM Tris-HCl, pH 7.5, 1% Nonidet P40, 100 mM KCl, 10 mM MgCl2 , 1 mM dithiothreitol), containing a protease inhibitor cocktail (Complete; Roche, Mannheim, Germany). ..

Article Title: LZAP Inhibits p38 MAPK (p38) Phosphorylation and Activity by Facilitating p38 Association with the Wild-Type p53 Induced Phosphatase 1 (WIP1)
Article Snippet: .. Immunoprecipitation and immunoblotting Cells were lysed in 0.5%(v/v) Nonidet P40 lysis buffer supplemented with protease inhibitor cocktail (Roche). .. Total cell extracts were incubated with specific antibodies and precipitated with protein A or G sepharose beads (GE Healthcare) before washing and suspension in Laemmli and gel electrophoresis followed by immunoblotting as described

Article Title: RASSF1A uncouples Wnt from Hippo signalling and promotes YAP mediated differentiation via p73
Article Snippet: .. For lysate preparation, 7 × 106 cells were lysed in ice-cold 1% (v/v) Nonidet P40 lysis buffer (20 mM Hepes pH 7.5, 1% NP40 (Roche), 150 mM NaCl, 0.5 mM EDTA, 50 mM NaF, 10 mM β-glycerophosphate, 0.5 mM Na3 VO4 and 1 × EDTA free protease inhibitors (Roche)). ..

Article Title: Molecular interactions of Bcl-2 and Bcl-xL with mortalin: identification and functional characterization
Article Snippet: .. Cell lysates were prepared in Nonidet P40 lysis buffer [10 mM Tris/HCl (pH 7.4), 150 mM NaCl, 5 mM EDTA, 1% Nonidet P40] supplemented with a protease inhibitor cocktail (Complete™ Mini; Roche Diagnostics) following 48 h of transfection. .. Stable cells lines were obtained by 2 weeks of selection for the transfected gene using 800 μg/ml Gly418 (Invitrogen) in the medium, and then maintained with 200 μg/ml Gly418 in DMEM.

Article Title: The non-structural protein μNS of piscine orthoreovirus (PRV) forms viral factory-like structures
Article Snippet: .. Cells were collected from the culture flasks 72 h post transfection (hpt), centrifuged at 5000 g for 5 min and resuspended in 1 mL Nonidet-P40 lysis buffer (1% NP-40, 50 mM Tris–HCl pH 8.0, 150 mM NaCl, 2 mM EDTA) containing Complete ultra mini protease inhibitor cocktail (Roche, Mannheim, Germany). .. The supernatant was transferred to a new tube, added antibodies against the desired epitope tag or anti-S8ORF2 and incubated overnight at 4 °C with rotation.

Article Title: Viral Protein Kinetics of Piscine Orthoreovirus Infection in Atlantic Salmon Blood Cells
Article Snippet: .. The cell pellets were lysed in Nonidet-P40 lysis buffer (1% NP-40, 50 mM Tris–HCl pH 8.0, 150 mM NaCl, 2 mM EDTA) containing Complete ultra mini protease inhibitor cocktail (Roche, Mannheim, Germany). .. The supernatant was mixed with Sample Buffer (Bio-Rad) and Reducing Agent (Bio-Rad), denatured for 5 min at 95 °C and run in SDS-PAGE, using 4%–12% Bis–Tris Criterion XT gel (Bio-Rad).

Article Title: RASSF1A uncouples Wnt from Hippo signalling and promotes YAP mediated differentiation via p73
Article Snippet: .. Co-immunoprecipitation For lysate preparation, 7 × 106 cells were lysed in ice-cold 1% (v/v) Nonidet P40 lysis buffer (20 mM Hepes pH 7.5, 1% NP40 (Roche), 150 mM NaCl, 0.5 mM EDTA, 50 mM NaF, 10 mM β-glycerophosphate, 0.5 mM Na3 VO4 and 1 × EDTA free protease inhibitors (Roche)). ..

Article Title: Wip1 phosphatase modulates both long-term potentiation and long-term depression through the dephosphorylation of CaMKII
Article Snippet: .. Tissues were lysed in 0.5% (v/v) Nonidet P40 lysis buffer supplemented with a protease inhibitor cocktail and PhosStop (Roche). .. The total cell extracts were incubated with specific antibodies and precipitated with protein A/G sepharose beads (sigma) before washing and suspension in Laemmli.

SDS Page:

Article Title: WDRPUH, A Novel WD-Repeat-Containing Protein, Is Highly Expressed in Human Hepatocellular Carcinoma and Involved in Cell Proliferation 1WDRPUH, A Novel WD-Repeat-Containing Protein, Is Highly Expressed in Human Hepatocellular Carcinoma and Involved in Cell Proliferation 1 *
Article Snippet: The cells were harvested at 48 hours after transfection and lysed with Nonidet P40 lysis buffer [20 mM Tris-HCl pH 7.5, 150 mM NaCl, 1% NP-40, 1x complete Protease Inhibitor Cocktail EDTA (-) (Roche, Basel, Switzerland)]. .. The eluted complex was centrifuged and supernatant was resuspended in 5x Laemmli sample buffer and separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE).

Article Title: Erythroid Progenitor Cells in Atlantic Salmon (Salmo salar) May Be Persistently and Productively Infected with Piscine Orthoreovirus (PRV)
Article Snippet: For each sample, 15 µL blood pellet was added to Nonidet-P40 lysis buffer containing complete ultra mini protease inhibitor cocktail (1:5) (Roche, Mannheim, Germany) and incubated for 30 min on ice. .. After centrifugation at 5000× g for 5 min, the supernatant was diluted with DEPC treated water (1:5) and mixed with XT sample buffer and XT reducing agent (Bio-Rad, Hercules, CA, USA) and boiled at 95 °C for 5 min. PRV proteins were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), using 4–12% criterion XT bis-tris gel.

Article Title: LPS-induced IL-8 activation in human intestinal epithelial cells is accompanied by specific histone H3 acetylation and methylation changes
Article Snippet: Western Blot Analysis Cell extracts were prepared in Nonidet P40 lysis buffer with 1 mM PMSF and Complete™ protease inhibitors mix (Roche, Indianapolis, IN, USA). .. 50 μg of proteins were resolved by electrophoresis using 10% SDS-PAGE gels and transferred to BA 85 0.45 μm PROTAN nitrocellulose filters (Schleicher & Schnell, Inc., Dassel, Germany).

Article Title: Caspase-mediated cleavage of the exosome subunit PM/Scl-75 during apoptosis
Article Snippet: Western blot analysis Cells were lysed on ice for 30 minutes in Nonidet P40 lysis buffer (25 mM Tris-HCl, pH 7.5, 1% Nonidet P40, 100 mM KCl, 10 mM MgCl2 , 1 mM dithiothreitol), containing a protease inhibitor cocktail (Complete; Roche, Mannheim, Germany). .. Protein extracts of 106 cells were analyzed by 10% SDS-PAGE and Western blotting with systemic lupus erythematosus patient serum Ven96 (a patient serum reactive with many exosome subunits), an anti-PM/Scl-100 rabbit serum [ ] or an anti-PM/Scl-75 rabbit or mouse serum [ ], followed by detection by means of horseradish peroxidase-conjugated secondary antibodies and visualization by chemiluminescence.

Article Title: Viral Protein Kinetics of Piscine Orthoreovirus Infection in Atlantic Salmon Blood Cells
Article Snippet: The cell pellets were lysed in Nonidet-P40 lysis buffer (1% NP-40, 50 mM Tris–HCl pH 8.0, 150 mM NaCl, 2 mM EDTA) containing Complete ultra mini protease inhibitor cocktail (Roche, Mannheim, Germany). .. The supernatant was mixed with Sample Buffer (Bio-Rad) and Reducing Agent (Bio-Rad), denatured for 5 min at 95 °C and run in SDS-PAGE, using 4%–12% Bis–Tris Criterion XT gel (Bio-Rad).

Plasmid Preparation:

Article Title: WDRPUH, A Novel WD-Repeat-Containing Protein, Is Highly Expressed in Human Hepatocellular Carcinoma and Involved in Cell Proliferation 1WDRPUH, A Novel WD-Repeat-Containing Protein, Is Highly Expressed in Human Hepatocellular Carcinoma and Involved in Cell Proliferation 1 *
Article Snippet: HEK293 cells that do not express WDRPUH were transfected with p3xFLAG-WDRPUH, mock vector, or p3xFLAG-SMYD3 (as a control). .. The cells were harvested at 48 hours after transfection and lysed with Nonidet P40 lysis buffer [20 mM Tris-HCl pH 7.5, 150 mM NaCl, 1% NP-40, 1x complete Protease Inhibitor Cocktail EDTA (-) (Roche, Basel, Switzerland)].

Article Title: The non-structural protein μNS of piscine orthoreovirus (PRV) forms viral factory-like structures
Article Snippet: Immunoprecipitation A total of 5 million EPC cells were pelleted by centrifugation, resuspended in 100 μL Ingenio Electroporation Solution (Mirus, Madison, WI, USA) and co-transfected with 8 μg plasmid using the Amaxa T-20 program. pcDNA3.1-μNS-N-FLAG was co-transfected with pcDNA3.1-σNS-N-MYC, pcDNA3.1-μ2-C-HA, pcDNA3.1-λ1-N-HA and pcDNA3.1 S8ORF2 (negative control) separately, using three parallel preparations. .. Cells were collected from the culture flasks 72 h post transfection (hpt), centrifuged at 5000 g for 5 min and resuspended in 1 mL Nonidet-P40 lysis buffer (1% NP-40, 50 mM Tris–HCl pH 8.0, 150 mM NaCl, 2 mM EDTA) containing Complete ultra mini protease inhibitor cocktail (Roche, Mannheim, Germany).

Negative Control:

Article Title: The non-structural protein μNS of piscine orthoreovirus (PRV) forms viral factory-like structures
Article Snippet: Immunoprecipitation A total of 5 million EPC cells were pelleted by centrifugation, resuspended in 100 μL Ingenio Electroporation Solution (Mirus, Madison, WI, USA) and co-transfected with 8 μg plasmid using the Amaxa T-20 program. pcDNA3.1-μNS-N-FLAG was co-transfected with pcDNA3.1-σNS-N-MYC, pcDNA3.1-μ2-C-HA, pcDNA3.1-λ1-N-HA and pcDNA3.1 S8ORF2 (negative control) separately, using three parallel preparations. .. Cells were collected from the culture flasks 72 h post transfection (hpt), centrifuged at 5000 g for 5 min and resuspended in 1 mL Nonidet-P40 lysis buffer (1% NP-40, 50 mM Tris–HCl pH 8.0, 150 mM NaCl, 2 mM EDTA) containing Complete ultra mini protease inhibitor cocktail (Roche, Mannheim, Germany).

Selection:

Article Title: Molecular interactions of Bcl-2 and Bcl-xL with mortalin: identification and functional characterization
Article Snippet: Cell lysates were prepared in Nonidet P40 lysis buffer [10 mM Tris/HCl (pH 7.4), 150 mM NaCl, 5 mM EDTA, 1% Nonidet P40] supplemented with a protease inhibitor cocktail (Complete™ Mini; Roche Diagnostics) following 48 h of transfection. .. Stable cells lines were obtained by 2 weeks of selection for the transfected gene using 800 μg/ml Gly418 (Invitrogen) in the medium, and then maintained with 200 μg/ml Gly418 in DMEM.

Immunoprecipitation:

Article Title: WDRPUH, A Novel WD-Repeat-Containing Protein, Is Highly Expressed in Human Hepatocellular Carcinoma and Involved in Cell Proliferation 1WDRPUH, A Novel WD-Repeat-Containing Protein, Is Highly Expressed in Human Hepatocellular Carcinoma and Involved in Cell Proliferation 1 *
Article Snippet: The cells were harvested at 48 hours after transfection and lysed with Nonidet P40 lysis buffer [20 mM Tris-HCl pH 7.5, 150 mM NaCl, 1% NP-40, 1x complete Protease Inhibitor Cocktail EDTA (-) (Roche, Basel, Switzerland)]. .. Cell lysates were immunoprecipitated with anti-FLAG M2 antibody conjugated with agarose overnight at 4°C.

Article Title: LZAP Inhibits p38 MAPK (p38) Phosphorylation and Activity by Facilitating p38 Association with the Wild-Type p53 Induced Phosphatase 1 (WIP1)
Article Snippet: .. Immunoprecipitation and immunoblotting Cells were lysed in 0.5%(v/v) Nonidet P40 lysis buffer supplemented with protease inhibitor cocktail (Roche). .. Total cell extracts were incubated with specific antibodies and precipitated with protein A or G sepharose beads (GE Healthcare) before washing and suspension in Laemmli and gel electrophoresis followed by immunoblotting as described

Article Title: The non-structural protein μNS of piscine orthoreovirus (PRV) forms viral factory-like structures
Article Snippet: Paragraph title: Immunoprecipitation ... Cells were collected from the culture flasks 72 h post transfection (hpt), centrifuged at 5000 g for 5 min and resuspended in 1 mL Nonidet-P40 lysis buffer (1% NP-40, 50 mM Tris–HCl pH 8.0, 150 mM NaCl, 2 mM EDTA) containing Complete ultra mini protease inhibitor cocktail (Roche, Mannheim, Germany).

Article Title: Wip1 phosphatase modulates both long-term potentiation and long-term depression through the dephosphorylation of CaMKII
Article Snippet: Paragraph title: Immunoprecipitation ... Tissues were lysed in 0.5% (v/v) Nonidet P40 lysis buffer supplemented with a protease inhibitor cocktail and PhosStop (Roche).

Fractionation:

Article Title: Endogenous RGS14 is a cytoplasmic-nuclear shuttling protein that localizes to juxtanuclear membranes and chromatin-rich regions of the nucleus
Article Snippet: Paragraph title: Subcellular fractionation ... Cells were then resuspended in 10 volumes of Nonidet-P40 lysis buffer (10 mM HEPES, pH 7.5; 10 mM KCl; 0.1 mM EDTA; 1 mM dithiothreitol (DTT); 0.5% Nonidet‐40; protease inhibitor cocktail (Roche)) and allowed to swell in ice for 12 min with intermittent mixing.

Marker:

Article Title: Viral Protein Kinetics of Piscine Orthoreovirus Infection in Atlantic Salmon Blood Cells
Article Snippet: The cell pellets were lysed in Nonidet-P40 lysis buffer (1% NP-40, 50 mM Tris–HCl pH 8.0, 150 mM NaCl, 2 mM EDTA) containing Complete ultra mini protease inhibitor cocktail (Roche, Mannheim, Germany). .. Magic MarkTM XP Standard (Invitrogen) was used as a molecular size marker.

Staining:

Article Title: LPS-induced IL-8 activation in human intestinal epithelial cells is accompanied by specific histone H3 acetylation and methylation changes
Article Snippet: Western Blot Analysis Cell extracts were prepared in Nonidet P40 lysis buffer with 1 mM PMSF and Complete™ protease inhibitors mix (Roche, Indianapolis, IN, USA). .. Immunoblots were stained with correspondent secondary antibodies IgG (Amersham Pharmacia Biotech, Buckinghamshire, UK), and revealed with the enhanced chemiluminescence detection system IgG (ECL, Amersham Pharmacia Biotech, Buckinghamshire, UK).

Article Title: Viral Protein Kinetics of Piscine Orthoreovirus Infection in Atlantic Salmon Blood Cells
Article Snippet: Antisera against µNS (1:1000) and λ1 (1:500); secondary antibody against rabbit IgG conjugated with Alexa Fluor 488 (Life Technologies) and Hoechst trihydrochloride trihydrate (Life Technologies) were used for staining. .. The cell pellets were lysed in Nonidet-P40 lysis buffer (1% NP-40, 50 mM Tris–HCl pH 8.0, 150 mM NaCl, 2 mM EDTA) containing Complete ultra mini protease inhibitor cocktail (Roche, Mannheim, Germany).

Fluorescence In Situ Hybridization:

Article Title: Erythroid Progenitor Cells in Atlantic Salmon (Salmo salar) May Be Persistently and Productively Infected with Piscine Orthoreovirus (PRV)
Article Snippet: Heparinized blood from PRV-1 infected and uninfected fish from a previous challenge trial were used as positive and negative controls, respectively [ ]. .. For each sample, 15 µL blood pellet was added to Nonidet-P40 lysis buffer containing complete ultra mini protease inhibitor cocktail (1:5) (Roche, Mannheim, Germany) and incubated for 30 min on ice.

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