protease inhibitor cocktail  (Roche)


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    Structured Review

    Roche protease inhibitor cocktail
    Protease Inhibitor Cocktail, supplied by Roche, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/protease inhibitor cocktail/product/Roche
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    protease inhibitor cocktail - by Bioz Stars, 2021-03
    86/100 stars

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    Related Articles

    Purification:

    Article Title: Preparation of Protein Samples for NMR Structure, Function, and Small Molecule Screening Studies
    Article Snippet: The database also reports associated protein target characteristics critical for the purification effort including protein molecular weight with tag for the exact isotope enrichment strategy, protein isoelectric point and extinction coefficient among others. .. The centrifuge bags of targets selected for purification are thawed on ice, 30 mL of Binding Buffer (50 mM Tris-HCl, 500 mM NaCl, 40 mM Imidazole, 1 mM TCEP and 0.02% NaN3 , pH 7.5) containing a protease inhibitor cocktail (Complete Protease Inhibitor Cocktail, Roche) is pipetted into each bag and the pellet is resuspended by simple hand manipulation of the cell paste. .. The bag contents are then transferred to a pre-chilled stainless steel 125 mL beaker (Vollrath®) and sonicated in an ice water bath using a Dual Horn ¾” probe (Qsonica, LLC.) for ten minutes with 30 second bursts followed by 30 second cooling periods (30 seconds on/30 seconds off).

    Binding Assay:

    Article Title: Preparation of Protein Samples for NMR Structure, Function, and Small Molecule Screening Studies
    Article Snippet: The database also reports associated protein target characteristics critical for the purification effort including protein molecular weight with tag for the exact isotope enrichment strategy, protein isoelectric point and extinction coefficient among others. .. The centrifuge bags of targets selected for purification are thawed on ice, 30 mL of Binding Buffer (50 mM Tris-HCl, 500 mM NaCl, 40 mM Imidazole, 1 mM TCEP and 0.02% NaN3 , pH 7.5) containing a protease inhibitor cocktail (Complete Protease Inhibitor Cocktail, Roche) is pipetted into each bag and the pellet is resuspended by simple hand manipulation of the cell paste. .. The bag contents are then transferred to a pre-chilled stainless steel 125 mL beaker (Vollrath®) and sonicated in an ice water bath using a Dual Horn ¾” probe (Qsonica, LLC.) for ten minutes with 30 second bursts followed by 30 second cooling periods (30 seconds on/30 seconds off).

    Protease Inhibitor:

    Article Title: Preparation of Protein Samples for NMR Structure, Function, and Small Molecule Screening Studies
    Article Snippet: The database also reports associated protein target characteristics critical for the purification effort including protein molecular weight with tag for the exact isotope enrichment strategy, protein isoelectric point and extinction coefficient among others. .. The centrifuge bags of targets selected for purification are thawed on ice, 30 mL of Binding Buffer (50 mM Tris-HCl, 500 mM NaCl, 40 mM Imidazole, 1 mM TCEP and 0.02% NaN3 , pH 7.5) containing a protease inhibitor cocktail (Complete Protease Inhibitor Cocktail, Roche) is pipetted into each bag and the pellet is resuspended by simple hand manipulation of the cell paste. .. The bag contents are then transferred to a pre-chilled stainless steel 125 mL beaker (Vollrath®) and sonicated in an ice water bath using a Dual Horn ¾” probe (Qsonica, LLC.) for ten minutes with 30 second bursts followed by 30 second cooling periods (30 seconds on/30 seconds off).

    Article Title: Engineering of Bacillus thuringiensis Cry Proteins to Enhance the Activity against Western Corn Rootworm
    Article Snippet: The shuffled IP3 protein was diluted to 0.5 mg/mL in phosphate buffer saline containing 20% of the midgut fluid and incubated at 25 °C to observe the digestion in vitro. .. Two μL of this mixture were taken at predetermined time intervals and mixed with 20 μL of NuPAGE® LDS sample buffer (Life Technologies, Carlsbad, CA, USA) that contained a reducing agent and Roche cOmplete™ Protease Inhibitor Cocktail obtained from Sigma-Aldrich (St Louis, MO, USA) at 2.5-fold higher concentration than the manufacturer’s suggested dilution. .. The samples mixed with LDS were heated immediately at 100 °C for 5 min and then analyzed by SDS-PAGE using NuPAGE® 4-12% Bis-Tris precast gel (Life Technologies, Carlsbad, CA, USA).

    Article Title: Computational modeling suggests dimerization of equine infectious anemia virus Rev is required for RNA binding
    Article Snippet: Expression and purification of EIAV Rev MBP-Rev fusion proteins were cloned and expressed in E. coli strain Rosetta Gami in NZY media as described previously [ ]. .. Following expression, cells were pelleted and resuspended in lysis buffer containing 25 mM HEPES pH 7.5, 200 mM NaCl, 2 mM beta-mercaptoethanol (BME), supplemented with 2 mM phenylmethylsulfonyl fluoride (PMSF) and Roche cOmplete® protease inhibitor cocktail tablet, according to the manufacturer’s protocol. .. The suspension was incubated with 1 mg/ml lysozyme on ice for 20 min and subjected to 10 cycles of freeze-thaw and 20 cycles of sonication.

    Article Title: Oncolysis with DTT-205 and DTT-304 generates immunological memory in cured animals
    Article Snippet: Released HMGB1 protein content was quantified in supernatants by ELISA (#ST51011; IBL, Hamburg, Germany) according to the manufacturer’s recommendations, and samples were measured with a Paradigm E3 multilabel reader (Molecular Devices). .. Western blot Half a million cells were harvested from six-well plates resuspended in lysis buffer containing 150 mM sodium chloride, 1.0% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, and protease inhibitor cocktails (Complete protease inhibitor cocktail, Roche, Basel, Switzerland) and incubated on ice for 30 min. To obtain supernatant, cell lysate was centrifuged at 12000×g for 20 min at 4 °C to remove insoluble materials. .. The lysate was mixed with 4 × NuPAGE® LDS sample buffer and 10 × sample reducing agent, and proteins were denaturated at 100℃ for 10 min. NuPAGE® Novex® 4–12% Bis-Tris Protein Gels (Thermo Fisher Scientific) were used for protein electrophoresis under a 100 V constant voltage mode.

    Article Title: Humanizing the yeast origin recognition complex
    Article Snippet: In brief, WT and mutant yeast cell pellets were incubated on ice for 3 min with 50 µl of cold ATAC-RSB (10 mM Tris HCl (pH 7.4), 10 mM NaCl, 3 mM MgCl2 , 1× ROCHE Complete Protease Inhibitor Cocktail, 0.1% IGEPAL CA-630, 0.1% Tween-20, and 0.01% digitonin). .. Subsequently, 1 ml of cold ATAC-RSB with 0.1% Tween-20 (10 mM Tris HCl (pH 7.4), 10 mM NaCl, 3 mM MgCl2, 1× ROCHE Complete Protease Inhibitor Cocktail, and 0.1% Tween-20) was added to the mixture. .. Nuclei were recovered by centrifugation and resuspended in transposase mix for tagmentation (1× TTBL and 2.5 μl TTE Mix V50 (Vazyme TD501 TruePrep DNA Library Prep Kit V2 for Illumina®), 0.01% digitonin, 0.1% Tween-20, and PBS).

    Article Title: The proteome of cytosolic lipid droplets isolated from differentiated Caco-2/TC7 enterocytes reveals cell-specific characteristics
    Article Snippet: Subcellular fractionation Lipid droplets were isolated by density-gradient centrifugation from Caco-2/TC7 cells incubated for 24 h with lipid micelles, using a protocol adapted from Yu et al. ( ) and Brasaemle and Wolins ( ). .. Cell layers (two 75 mm diameter inserts per centrifuge tube) were rinsed briefly (three times with cold PBS) and scraped and the volume of the cell homogenate was adjusted to 2 ml with buffer A (25 mM Tris/HCl, pH 7.4, 100 mM KCl, 1 mM EDTA and 5 mM EGTA) containing a protease inhibitor cocktail (Complete® protease inhibitor cocktail from Roche). .. Cells were lysed twice using a cell disruption bomb (Parr Instrument Company; 1200 lbf/in2 , 10 min; 1 lbf/in2 =6.9 kPa).

    Article Title: Oesophagostomum dentatum Extract Modulates T Cell-Dependent Immune Responses to Bystander Antigens and Prevents the Development of Allergy in Mice
    Article Snippet: Mice were terminally anesthetized 72 h after the final airway challenge (day 40). .. Bronchoalvelar Lavage and Differential Cell Counts Bronchoalveolar lavage fluid (BALF) was collected by cannulating the trachea and injecting/recovering 2×1 ml PBS containing a protease inhibitor cocktail (Complete® Protease Inhibitor cocktail Tablets; Roche, Mannheim, Germany). ..

    Concentration Assay:

    Article Title: Engineering of Bacillus thuringiensis Cry Proteins to Enhance the Activity against Western Corn Rootworm
    Article Snippet: The shuffled IP3 protein was diluted to 0.5 mg/mL in phosphate buffer saline containing 20% of the midgut fluid and incubated at 25 °C to observe the digestion in vitro. .. Two μL of this mixture were taken at predetermined time intervals and mixed with 20 μL of NuPAGE® LDS sample buffer (Life Technologies, Carlsbad, CA, USA) that contained a reducing agent and Roche cOmplete™ Protease Inhibitor Cocktail obtained from Sigma-Aldrich (St Louis, MO, USA) at 2.5-fold higher concentration than the manufacturer’s suggested dilution. .. The samples mixed with LDS were heated immediately at 100 °C for 5 min and then analyzed by SDS-PAGE using NuPAGE® 4-12% Bis-Tris precast gel (Life Technologies, Carlsbad, CA, USA).

    Expressing:

    Article Title: Computational modeling suggests dimerization of equine infectious anemia virus Rev is required for RNA binding
    Article Snippet: Expression and purification of EIAV Rev MBP-Rev fusion proteins were cloned and expressed in E. coli strain Rosetta Gami in NZY media as described previously [ ]. .. Following expression, cells were pelleted and resuspended in lysis buffer containing 25 mM HEPES pH 7.5, 200 mM NaCl, 2 mM beta-mercaptoethanol (BME), supplemented with 2 mM phenylmethylsulfonyl fluoride (PMSF) and Roche cOmplete® protease inhibitor cocktail tablet, according to the manufacturer’s protocol. .. The suspension was incubated with 1 mg/ml lysozyme on ice for 20 min and subjected to 10 cycles of freeze-thaw and 20 cycles of sonication.

    Lysis:

    Article Title: Computational modeling suggests dimerization of equine infectious anemia virus Rev is required for RNA binding
    Article Snippet: Expression and purification of EIAV Rev MBP-Rev fusion proteins were cloned and expressed in E. coli strain Rosetta Gami in NZY media as described previously [ ]. .. Following expression, cells were pelleted and resuspended in lysis buffer containing 25 mM HEPES pH 7.5, 200 mM NaCl, 2 mM beta-mercaptoethanol (BME), supplemented with 2 mM phenylmethylsulfonyl fluoride (PMSF) and Roche cOmplete® protease inhibitor cocktail tablet, according to the manufacturer’s protocol. .. The suspension was incubated with 1 mg/ml lysozyme on ice for 20 min and subjected to 10 cycles of freeze-thaw and 20 cycles of sonication.

    Article Title: Oncolysis with DTT-205 and DTT-304 generates immunological memory in cured animals
    Article Snippet: Released HMGB1 protein content was quantified in supernatants by ELISA (#ST51011; IBL, Hamburg, Germany) according to the manufacturer’s recommendations, and samples were measured with a Paradigm E3 multilabel reader (Molecular Devices). .. Western blot Half a million cells were harvested from six-well plates resuspended in lysis buffer containing 150 mM sodium chloride, 1.0% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, and protease inhibitor cocktails (Complete protease inhibitor cocktail, Roche, Basel, Switzerland) and incubated on ice for 30 min. To obtain supernatant, cell lysate was centrifuged at 12000×g for 20 min at 4 °C to remove insoluble materials. .. The lysate was mixed with 4 × NuPAGE® LDS sample buffer and 10 × sample reducing agent, and proteins were denaturated at 100℃ for 10 min. NuPAGE® Novex® 4–12% Bis-Tris Protein Gels (Thermo Fisher Scientific) were used for protein electrophoresis under a 100 V constant voltage mode.

    Western Blot:

    Article Title: Oncolysis with DTT-205 and DTT-304 generates immunological memory in cured animals
    Article Snippet: Released HMGB1 protein content was quantified in supernatants by ELISA (#ST51011; IBL, Hamburg, Germany) according to the manufacturer’s recommendations, and samples were measured with a Paradigm E3 multilabel reader (Molecular Devices). .. Western blot Half a million cells were harvested from six-well plates resuspended in lysis buffer containing 150 mM sodium chloride, 1.0% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, and protease inhibitor cocktails (Complete protease inhibitor cocktail, Roche, Basel, Switzerland) and incubated on ice for 30 min. To obtain supernatant, cell lysate was centrifuged at 12000×g for 20 min at 4 °C to remove insoluble materials. .. The lysate was mixed with 4 × NuPAGE® LDS sample buffer and 10 × sample reducing agent, and proteins were denaturated at 100℃ for 10 min. NuPAGE® Novex® 4–12% Bis-Tris Protein Gels (Thermo Fisher Scientific) were used for protein electrophoresis under a 100 V constant voltage mode.

    Incubation:

    Article Title: Oncolysis with DTT-205 and DTT-304 generates immunological memory in cured animals
    Article Snippet: Released HMGB1 protein content was quantified in supernatants by ELISA (#ST51011; IBL, Hamburg, Germany) according to the manufacturer’s recommendations, and samples were measured with a Paradigm E3 multilabel reader (Molecular Devices). .. Western blot Half a million cells were harvested from six-well plates resuspended in lysis buffer containing 150 mM sodium chloride, 1.0% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, and protease inhibitor cocktails (Complete protease inhibitor cocktail, Roche, Basel, Switzerland) and incubated on ice for 30 min. To obtain supernatant, cell lysate was centrifuged at 12000×g for 20 min at 4 °C to remove insoluble materials. .. The lysate was mixed with 4 × NuPAGE® LDS sample buffer and 10 × sample reducing agent, and proteins were denaturated at 100℃ for 10 min. NuPAGE® Novex® 4–12% Bis-Tris Protein Gels (Thermo Fisher Scientific) were used for protein electrophoresis under a 100 V constant voltage mode.

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    Roche immunoprecipitation buffer
    CNTNAP2 interacts with CASK at the plasma membrane in cortical GABAergic interneurons (a) Only yeast cells co-expressing CNTNAP2 bait and CASK prey constructs grow on high stringency yeast plates (QDO/X/A). (b–c) Cropped western blots of <t>co-immunoprecipitation</t> experiments with CASK and CNTNAP2 in mouse cortex. (d) Cropped western blots showing co-immunoprecipitation of various FLAG-CNTNAP2 truncation mutants ( Supplementary Figure 5a ; red lines) with untagged, full-length CASK in HEK293T cells. (e) Representative cropped western blots of membrane/cytosol fractions of HEK293T cells expressing pCS2-FLAG + CASK-mCherry (CASK), FLAG-CNTNAP2 + CASK-mCherry (CNTNAP2 + CASK), or FLAG-CNTNAP2 + CASKΔPDZ-mCherry (CNTNAP2 + CASKΔPDZ) and subsequent quantification of protein localization (CASK vs. CNTNAP2 + CASK vs. CNTNAP2 + CASKΔPDZ: 6 independent experiments; CASK alone vs. CASKΔPDZ alone: 3 independent experiments). Percentages were calculated by dividing the densitometry value of CASK/CNTNAP2 in either membrane or cytosol fraction by the summation of both. (f) Representative SIM image of endogenous CNTNAP2 and CASK co-localization (white) on a GFP-transfected interneuronal dendrite (scale bar = 5 μm). (g) Histogram showing distribution of CASK/CNTNAP2 co-localized puncta relative to the dendrite’s lateral edge from (f) (CASK colocalized with CNTNAP2: n = 79 puncta from 3 cultures; CNTNAP2 colocalized with CASK: n = 70 puncta from 3 cultures). (h) Representative confocal image showing PLA signal from endogenous CASK/CNTNAP2 staining, which occurs only when CASK and CNTNAP2 primary antibodies are both applied (scale bar = 1 μm). (i) Cropped immunoblots of subcellular fractionations from adult mouse forebrain probed with CNTNAP2, CASK, and β-tubulin. CNTNAP2 and CASK, but not β-tubulin, are found in the washed membrane fraction (S5; red box). (j) Cropped western blot of time course and (k) quantification of CASK expression in cultured cortical neurons (n = 3 independent experiments). Values are means ± SEM. * P≤0.05, ** P≤0.01, *** P≤0.001; one-way ANOVA with Bonferroni’s correction (k, top graph; e). Student’s t-test (middle and bottom graphs; e).
    Immunoprecipitation Buffer, supplied by Roche, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/immunoprecipitation buffer/product/Roche
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    86
    Roche complete mini protease inhibitor cocktail
    Expression of conventional breast cancer biomarkers and selected multidrug resistance genes. Cell lysates were prepared in radioimmunoprecipitation assay buffer supplemented with <t>cOmplete</t> Mini protease inhibitor and PhosSTOP phosphatase inhibitor. Quantities (10–50 μg) of total protein were run on a 10 % gel (MDR1), 4–20 % precast gels (epidermal growth factor receptor [EGFR], oestrogen receptor [ER], progesterone receptor [PR], type II topoisomerase [TOPO IIα]) and Any kD Mini-PROTEAN TGX Precast Protein Gels (human epidermal growth factor receptor 2 [HER2], HER3), transferred onto polyvinylidene fluoride membrane and developed using chemiluminescence substrate. Nat native, Epi-R epirubicin-resistant, GAPDH glyceraldehyde-3-phosphate dehydrogenase
    Complete Mini Protease Inhibitor Cocktail, supplied by Roche, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/complete mini protease inhibitor cocktail/product/Roche
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
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    86
    Roche pbs containing protease inhibitor cocktail
    ST398 CA-MRSA virulence in a mouse model. The seven ST398 CA-MRSA isolates were compared with closely related ST398 MSSA, all ST398 LA-MSSA obtained for this study, and representative Chinese ST5 and ST239 HA-MRSA strains by in vivo analysis of virulence. In addition, two USA300 CA-MRSA strains (SF8300, LAC) and the genome-sequenced LA-MRSA S0385 standard strain were used in the comparisons. Isolate selection for the MSSA and HA-MRSA groups was performed, as in a previous study [ 26 ], by selecting isolates whose average PSM production was similar to that of the entire group and thus reflects the average virulence potential. Notably, for the in vivo infection studies, the ST398 CA-MRSA, MSSA, and ST5/ST239 HA-MRSA isolates were obtained from the corresponding type of human infection (respiratory and skin infection, respectively). See Additional file 1 : Table S2 for isolates that were compared and their characteristics. a Mouse skin infection model. Mice received 10 7 live S. aureus or <t>PBS</t> alone in the right flank by subcutaneous injection. Skin abscesses (length × width) were measured 48 h after infection. b Mouse lung infection model. We pipetted 4 × 10 9 CFU/40 μl S. aureus into the nares of the anesthetized mice. All mice were euthanized 48 h after inoculation and CFU in the lung tissue were determined. a , b Three mice were infected with each strain; the shown symbols represent the average value obtained from three mice. c – f Inflammatory <t>cytokine</t> expression in ST398 CA-MRSA isolates in comparison. The inflammatory cytokines TNF-α and IL-6 were determined in skin ( c , d ) or lung tissue ( e , f ). a – f Statistical analysis is by one-way ANOVA with Dunnett’s post test versus the ST398 CA-MRSA group. * p
    Pbs Containing Protease Inhibitor Cocktail, supplied by Roche, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    CNTNAP2 interacts with CASK at the plasma membrane in cortical GABAergic interneurons (a) Only yeast cells co-expressing CNTNAP2 bait and CASK prey constructs grow on high stringency yeast plates (QDO/X/A). (b–c) Cropped western blots of co-immunoprecipitation experiments with CASK and CNTNAP2 in mouse cortex. (d) Cropped western blots showing co-immunoprecipitation of various FLAG-CNTNAP2 truncation mutants ( Supplementary Figure 5a ; red lines) with untagged, full-length CASK in HEK293T cells. (e) Representative cropped western blots of membrane/cytosol fractions of HEK293T cells expressing pCS2-FLAG + CASK-mCherry (CASK), FLAG-CNTNAP2 + CASK-mCherry (CNTNAP2 + CASK), or FLAG-CNTNAP2 + CASKΔPDZ-mCherry (CNTNAP2 + CASKΔPDZ) and subsequent quantification of protein localization (CASK vs. CNTNAP2 + CASK vs. CNTNAP2 + CASKΔPDZ: 6 independent experiments; CASK alone vs. CASKΔPDZ alone: 3 independent experiments). Percentages were calculated by dividing the densitometry value of CASK/CNTNAP2 in either membrane or cytosol fraction by the summation of both. (f) Representative SIM image of endogenous CNTNAP2 and CASK co-localization (white) on a GFP-transfected interneuronal dendrite (scale bar = 5 μm). (g) Histogram showing distribution of CASK/CNTNAP2 co-localized puncta relative to the dendrite’s lateral edge from (f) (CASK colocalized with CNTNAP2: n = 79 puncta from 3 cultures; CNTNAP2 colocalized with CASK: n = 70 puncta from 3 cultures). (h) Representative confocal image showing PLA signal from endogenous CASK/CNTNAP2 staining, which occurs only when CASK and CNTNAP2 primary antibodies are both applied (scale bar = 1 μm). (i) Cropped immunoblots of subcellular fractionations from adult mouse forebrain probed with CNTNAP2, CASK, and β-tubulin. CNTNAP2 and CASK, but not β-tubulin, are found in the washed membrane fraction (S5; red box). (j) Cropped western blot of time course and (k) quantification of CASK expression in cultured cortical neurons (n = 3 independent experiments). Values are means ± SEM. * P≤0.05, ** P≤0.01, *** P≤0.001; one-way ANOVA with Bonferroni’s correction (k, top graph; e). Student’s t-test (middle and bottom graphs; e).

    Journal: Molecular psychiatry

    Article Title: CNTNAP2 stabilizes interneuron dendritic arbors through CASK

    doi: 10.1038/s41380-018-0027-3

    Figure Lengend Snippet: CNTNAP2 interacts with CASK at the plasma membrane in cortical GABAergic interneurons (a) Only yeast cells co-expressing CNTNAP2 bait and CASK prey constructs grow on high stringency yeast plates (QDO/X/A). (b–c) Cropped western blots of co-immunoprecipitation experiments with CASK and CNTNAP2 in mouse cortex. (d) Cropped western blots showing co-immunoprecipitation of various FLAG-CNTNAP2 truncation mutants ( Supplementary Figure 5a ; red lines) with untagged, full-length CASK in HEK293T cells. (e) Representative cropped western blots of membrane/cytosol fractions of HEK293T cells expressing pCS2-FLAG + CASK-mCherry (CASK), FLAG-CNTNAP2 + CASK-mCherry (CNTNAP2 + CASK), or FLAG-CNTNAP2 + CASKΔPDZ-mCherry (CNTNAP2 + CASKΔPDZ) and subsequent quantification of protein localization (CASK vs. CNTNAP2 + CASK vs. CNTNAP2 + CASKΔPDZ: 6 independent experiments; CASK alone vs. CASKΔPDZ alone: 3 independent experiments). Percentages were calculated by dividing the densitometry value of CASK/CNTNAP2 in either membrane or cytosol fraction by the summation of both. (f) Representative SIM image of endogenous CNTNAP2 and CASK co-localization (white) on a GFP-transfected interneuronal dendrite (scale bar = 5 μm). (g) Histogram showing distribution of CASK/CNTNAP2 co-localized puncta relative to the dendrite’s lateral edge from (f) (CASK colocalized with CNTNAP2: n = 79 puncta from 3 cultures; CNTNAP2 colocalized with CASK: n = 70 puncta from 3 cultures). (h) Representative confocal image showing PLA signal from endogenous CASK/CNTNAP2 staining, which occurs only when CASK and CNTNAP2 primary antibodies are both applied (scale bar = 1 μm). (i) Cropped immunoblots of subcellular fractionations from adult mouse forebrain probed with CNTNAP2, CASK, and β-tubulin. CNTNAP2 and CASK, but not β-tubulin, are found in the washed membrane fraction (S5; red box). (j) Cropped western blot of time course and (k) quantification of CASK expression in cultured cortical neurons (n = 3 independent experiments). Values are means ± SEM. * P≤0.05, ** P≤0.01, *** P≤0.001; one-way ANOVA with Bonferroni’s correction (k, top graph; e). Student’s t-test (middle and bottom graphs; e).

    Article Snippet: Expression Time Course Cortices from CD1 WT mice aged P0, P14, P28, 4 months, and 6 months (sex not determined at P0, P14; males for P28, 4 months, 6 months) were homogenized in immunoprecipitation buffer (50 mM Tris pH 7.4, 150 mM NaCl, 0.5% Triton X-100 with Roche protease inhibitor cocktail) and solubilized for 1 hour at 4°C.

    Techniques: Expressing, Construct, Western Blot, Immunoprecipitation, Transfection, Proximity Ligation Assay, Staining, Cell Culture

    Expression of conventional breast cancer biomarkers and selected multidrug resistance genes. Cell lysates were prepared in radioimmunoprecipitation assay buffer supplemented with cOmplete Mini protease inhibitor and PhosSTOP phosphatase inhibitor. Quantities (10–50 μg) of total protein were run on a 10 % gel (MDR1), 4–20 % precast gels (epidermal growth factor receptor [EGFR], oestrogen receptor [ER], progesterone receptor [PR], type II topoisomerase [TOPO IIα]) and Any kD Mini-PROTEAN TGX Precast Protein Gels (human epidermal growth factor receptor 2 [HER2], HER3), transferred onto polyvinylidene fluoride membrane and developed using chemiluminescence substrate. Nat native, Epi-R epirubicin-resistant, GAPDH glyceraldehyde-3-phosphate dehydrogenase

    Journal: Breast Cancer Research : BCR

    Article Title: Downregulation of histone H2A and H2B pathways is associated with anthracycline sensitivity in breast cancer

    doi: 10.1186/s13058-016-0676-6

    Figure Lengend Snippet: Expression of conventional breast cancer biomarkers and selected multidrug resistance genes. Cell lysates were prepared in radioimmunoprecipitation assay buffer supplemented with cOmplete Mini protease inhibitor and PhosSTOP phosphatase inhibitor. Quantities (10–50 μg) of total protein were run on a 10 % gel (MDR1), 4–20 % precast gels (epidermal growth factor receptor [EGFR], oestrogen receptor [ER], progesterone receptor [PR], type II topoisomerase [TOPO IIα]) and Any kD Mini-PROTEAN TGX Precast Protein Gels (human epidermal growth factor receptor 2 [HER2], HER3), transferred onto polyvinylidene fluoride membrane and developed using chemiluminescence substrate. Nat native, Epi-R epirubicin-resistant, GAPDH glyceraldehyde-3-phosphate dehydrogenase

    Article Snippet: WCL were prepared by adding equal volumes of 2× RIPA buffer supplemented with 25 U of Benzonase Nuclease (Sigma-Aldrich) and cOmplete Mini Protease Inhibitor Cocktail (Roche), incubating on ice for 30 minutes and sonicating for 15 minutes with 30-second on-off intervals.

    Techniques: Expressing, Radio Immunoprecipitation, Protease Inhibitor

    ST398 CA-MRSA virulence in a mouse model. The seven ST398 CA-MRSA isolates were compared with closely related ST398 MSSA, all ST398 LA-MSSA obtained for this study, and representative Chinese ST5 and ST239 HA-MRSA strains by in vivo analysis of virulence. In addition, two USA300 CA-MRSA strains (SF8300, LAC) and the genome-sequenced LA-MRSA S0385 standard strain were used in the comparisons. Isolate selection for the MSSA and HA-MRSA groups was performed, as in a previous study [ 26 ], by selecting isolates whose average PSM production was similar to that of the entire group and thus reflects the average virulence potential. Notably, for the in vivo infection studies, the ST398 CA-MRSA, MSSA, and ST5/ST239 HA-MRSA isolates were obtained from the corresponding type of human infection (respiratory and skin infection, respectively). See Additional file 1 : Table S2 for isolates that were compared and their characteristics. a Mouse skin infection model. Mice received 10 7 live S. aureus or PBS alone in the right flank by subcutaneous injection. Skin abscesses (length × width) were measured 48 h after infection. b Mouse lung infection model. We pipetted 4 × 10 9 CFU/40 μl S. aureus into the nares of the anesthetized mice. All mice were euthanized 48 h after inoculation and CFU in the lung tissue were determined. a , b Three mice were infected with each strain; the shown symbols represent the average value obtained from three mice. c – f Inflammatory cytokine expression in ST398 CA-MRSA isolates in comparison. The inflammatory cytokines TNF-α and IL-6 were determined in skin ( c , d ) or lung tissue ( e , f ). a – f Statistical analysis is by one-way ANOVA with Dunnett’s post test versus the ST398 CA-MRSA group. * p

    Journal: Genome Medicine

    Article Title: Detection and analysis of methicillin-resistant human-adapted sequence type 398 allows insight into community-associated methicillin-resistant Staphylococcus aureus evolution

    doi: 10.1186/s13073-018-0514-9

    Figure Lengend Snippet: ST398 CA-MRSA virulence in a mouse model. The seven ST398 CA-MRSA isolates were compared with closely related ST398 MSSA, all ST398 LA-MSSA obtained for this study, and representative Chinese ST5 and ST239 HA-MRSA strains by in vivo analysis of virulence. In addition, two USA300 CA-MRSA strains (SF8300, LAC) and the genome-sequenced LA-MRSA S0385 standard strain were used in the comparisons. Isolate selection for the MSSA and HA-MRSA groups was performed, as in a previous study [ 26 ], by selecting isolates whose average PSM production was similar to that of the entire group and thus reflects the average virulence potential. Notably, for the in vivo infection studies, the ST398 CA-MRSA, MSSA, and ST5/ST239 HA-MRSA isolates were obtained from the corresponding type of human infection (respiratory and skin infection, respectively). See Additional file 1 : Table S2 for isolates that were compared and their characteristics. a Mouse skin infection model. Mice received 10 7 live S. aureus or PBS alone in the right flank by subcutaneous injection. Skin abscesses (length × width) were measured 48 h after infection. b Mouse lung infection model. We pipetted 4 × 10 9 CFU/40 μl S. aureus into the nares of the anesthetized mice. All mice were euthanized 48 h after inoculation and CFU in the lung tissue were determined. a , b Three mice were infected with each strain; the shown symbols represent the average value obtained from three mice. c – f Inflammatory cytokine expression in ST398 CA-MRSA isolates in comparison. The inflammatory cytokines TNF-α and IL-6 were determined in skin ( c , d ) or lung tissue ( e , f ). a – f Statistical analysis is by one-way ANOVA with Dunnett’s post test versus the ST398 CA-MRSA group. * p

    Article Snippet: Length (L) and width (W) values were used to calculate the area of lesions with the formula L × W. Mouse skin tissue of the same size was taken from the abscess and PBS control groups and homogenated with glass beads with PBS-containing protease inhibitor cocktail (Roche) for cytokine detection.

    Techniques: In Vivo, Selection, Infection, Mouse Assay, Injection, Expressing

    : Table S2 for isolates that were compared and their characteristics. a Mouse skin infection model. Mice received 10 7 live S. aureus or PBS alone in the right flank by subcutaneous injection. Skin abscesses (length × width) were measured 48 h after infection. b Mouse lung infection model. We pipetted 4 × 10 9 CFU/40 μl S. aureus into the nares of the anesthetized mice. All mice were euthanized 48 h after inoculation and CFU in the lung tissue were determined. a , b Three mice were infected with each strain; the shown symbols represent the average value obtained from three mice. c – f Inflammatory cytokine expression in ST398 CA-MRSA isolates in comparison. The inflammatory cytokines TNF-α and IL-6 were determined in skin ( c , d ) or lung tissue ( e , f ). a – f Statistical analysis is by one-way ANOVA with Dunnett’s post test versus the ST398 CA-MRSA group. * p

    Journal: Genome Medicine

    Article Title: Detection and analysis of methicillin-resistant human-adapted sequence type 398 allows insight into community-associated methicillin-resistant Staphylococcus aureus evolution

    doi: 10.1186/s13073-018-0514-9

    Figure Lengend Snippet: : Table S2 for isolates that were compared and their characteristics. a Mouse skin infection model. Mice received 10 7 live S. aureus or PBS alone in the right flank by subcutaneous injection. Skin abscesses (length × width) were measured 48 h after infection. b Mouse lung infection model. We pipetted 4 × 10 9 CFU/40 μl S. aureus into the nares of the anesthetized mice. All mice were euthanized 48 h after inoculation and CFU in the lung tissue were determined. a , b Three mice were infected with each strain; the shown symbols represent the average value obtained from three mice. c – f Inflammatory cytokine expression in ST398 CA-MRSA isolates in comparison. The inflammatory cytokines TNF-α and IL-6 were determined in skin ( c , d ) or lung tissue ( e , f ). a – f Statistical analysis is by one-way ANOVA with Dunnett’s post test versus the ST398 CA-MRSA group. * p

    Article Snippet: Length (L) and width (W) values were used to calculate the area of lesions with the formula L × W. Mouse skin tissue of the same size was taken from the abscess and PBS control groups and homogenated with glass beads with PBS-containing protease inhibitor cocktail (Roche) for cytokine detection.

    Techniques: Infection, Mouse Assay, Injection, Expressing