SDS Page:Article Title: A 3D Lymphatics-on-a-Chip Reveals Matrix Stiffness Dependent VEGFR-2 and VEGFR-3 Contributions to Lymphangiogenesis
Article Snippet: Treatment was performed 15 minutes before cold media were added to cells, which were then lysed in cold Pierce IP Lysis Buffer (ThermoFisher) with 2X cOmplete ULTRA Tablets, Mini, EASYpack Protease Inhibitor Cocktail (Sigma), and PhosStop (Sigma) phosphatase inhibitors, combined with 6X Laemmli SDS Sample Buffer (Boston Bioproducts) before boiling at 95°C for 10 minutes. .. The lysates, a PageRuler Plus Prestained Protein Ladder (ThermoFisher), and Precision Plus Protein Dual Color Standards (BioRad) were run in an SDS-PAGE using 4–20% Mini-PROTEAN TGX Precast Protein Gels (BioRad) and 1X Tris/Glycine SDS running buffer (BioRad) in a Mini-PROTEAN Tetra Cell (BioRad) at 120 V. Transfer to a PVDF membrane using a TransBlot Turbo RTA Mini 0.45 μm LF PVDF Transfer Kit (BioRad) and TransBlot Turbo Transfer Starter System (BioRad) was done at 2.5A and 25V. .. The membrane was rinsed in 1X TBST, then blocked in EveryBlot Blocking Buffer (BioRad) for 1 hour.
Article Title: Circular RNA vaccines encoding Pvs25 induce antibodies that block Plasmodium vivax transmission.
Article Snippet: Culture supernatants were collected, and cells were lysed in 1× Radioimmunoprecipitation (RIPA) buffer (Millipore 20–188) containing Protease Inhibitor Cocktail III (1:500, Millipore 539,134) and 1 mM Phenylmethanesulfonyl fluoride (PMSF) in isopropanol (Sigma-Aldrich 78,830). .. Proteins were separated SDS-PAGE under reducing condition on 4–20% Mini-PROTEAN TGX Precast Protein Gels (BioRad, Hercules, CA, USA) along with recombinant Pvs25 protein (10 ng) produced by the wheat germ cell-free system (WGCFS) as a positive control [16] and transferred to polyvinylidene fluoride (PVDF) membrane using a Trans-Blot Turbo Transfer System (BioRad) at 1.3A, 25 V for 7 min. Membranes were blocked with BlockPRO Protein-free Blocking Buffer (Energenesis Biomedical, Taipei, Taiwan) for 3 min and incubated with pooled antisera from mice immunized with recombinant Pvs25 (1:20,000) and a mouse anti-β-actin antibody (1:10,000; Abcam ab8226, Cambridge, UK) in 5% (w/v) skim milk in Tris-buffered saline containing Tween 20 (TBST). .. After three washes with TBST, the membranes were incubated with horseradish peroxidase (HRP)-conjugated anti-mouse IgG secondary antibody (1:10,000; Abcam ab6789).
Membrane:Article Title: A 3D Lymphatics-on-a-Chip Reveals Matrix Stiffness Dependent VEGFR-2 and VEGFR-3 Contributions to Lymphangiogenesis
Article Snippet: Treatment was performed 15 minutes before cold media were added to cells, which were then lysed in cold Pierce IP Lysis Buffer (ThermoFisher) with 2X cOmplete ULTRA Tablets, Mini, EASYpack Protease Inhibitor Cocktail (Sigma), and PhosStop (Sigma) phosphatase inhibitors, combined with 6X Laemmli SDS Sample Buffer (Boston Bioproducts) before boiling at 95°C for 10 minutes. .. The lysates, a PageRuler Plus Prestained Protein Ladder (ThermoFisher), and Precision Plus Protein Dual Color Standards (BioRad) were run in an SDS-PAGE using 4–20% Mini-PROTEAN TGX Precast Protein Gels (BioRad) and 1X Tris/Glycine SDS running buffer (BioRad) in a Mini-PROTEAN Tetra Cell (BioRad) at 120 V. Transfer to a PVDF membrane using a TransBlot Turbo RTA Mini 0.45 μm LF PVDF Transfer Kit (BioRad) and TransBlot Turbo Transfer Starter System (BioRad) was done at 2.5A and 25V. .. The membrane was rinsed in 1X TBST, then blocked in EveryBlot Blocking Buffer (BioRad) for 1 hour.
Article Title: Oligodendrocyte stress response as a driver of vascular dementia
Article Snippet: .. A total of 25 μg of protein was loaded onto a 4–20% Mini-PROTEAN TGX Precast Protein Gels and electrophoresed at 70 V for 15 min, followed by 120 V for 45 min. Dry transfer was performed on a PVDF membrane, previously activated with methanol, or a nitrocellulose membrane using a Trans-Blot Turbo Transfer System (Bio-Rad). ..
Article Title: Structural basis for start codon selection by eIF5-mimic proteins
Article Snippet: .. The total proteins in RRL samples were separated on 4 to 20% Mini-Protean TGX Precast Protein gels (Bio-Rad) alongside a protein marker (Bio-Rad) at 130 V for 2.5 hours in 1× tris-tricine-SDS running buffer, and proteins were transferred to a polyvinylidene difluoride membrane using a Trans-Blot Turbo transfer system (Bio-Rad). .. To reduce nonspecific signal, the membranes were blocked in blocking buffer [1× PBST containing 0.1% Tween 20 and 5% (v/v) nonfat dry milk (Research Products International, M17200 )] at room temperature for 30 min.
Article Title: Circular RNA vaccines encoding Pvs25 induce antibodies that block Plasmodium vivax transmission.
Article Snippet: Culture supernatants were collected, and cells were lysed in 1× Radioimmunoprecipitation (RIPA) buffer (Millipore 20–188) containing Protease Inhibitor Cocktail III (1:500, Millipore 539,134) and 1 mM Phenylmethanesulfonyl fluoride (PMSF) in isopropanol (Sigma-Aldrich 78,830). .. Proteins were separated SDS-PAGE under reducing condition on 4–20% Mini-PROTEAN TGX Precast Protein Gels (BioRad, Hercules, CA, USA) along with recombinant Pvs25 protein (10 ng) produced by the wheat germ cell-free system (WGCFS) as a positive control [16] and transferred to polyvinylidene fluoride (PVDF) membrane using a Trans-Blot Turbo Transfer System (BioRad) at 1.3A, 25 V for 7 min. Membranes were blocked with BlockPRO Protein-free Blocking Buffer (Energenesis Biomedical, Taipei, Taiwan) for 3 min and incubated with pooled antisera from mice immunized with recombinant Pvs25 (1:20,000) and a mouse anti-β-actin antibody (1:10,000; Abcam ab8226, Cambridge, UK) in 5% (w/v) skim milk in Tris-buffered saline containing Tween 20 (TBST). .. After three washes with TBST, the membranes were incubated with horseradish peroxidase (HRP)-conjugated anti-mouse IgG secondary antibody (1:10,000; Abcam ab6789).
Marker:Article Title: Structural basis for start codon selection by eIF5-mimic proteins
Article Snippet: .. The total proteins in RRL samples were separated on 4 to 20% Mini-Protean TGX Precast Protein gels (Bio-Rad) alongside a protein marker (Bio-Rad) at 130 V for 2.5 hours in 1× tris-tricine-SDS running buffer, and proteins were transferred to a polyvinylidene difluoride membrane using a Trans-Blot Turbo transfer system (Bio-Rad). .. To reduce nonspecific signal, the membranes were blocked in blocking buffer [1× PBST containing 0.1% Tween 20 and 5% (v/v) nonfat dry milk (Research Products International, M17200 )] at room temperature for 30 min.
Recombinant:Article Title: Circular RNA vaccines encoding Pvs25 induce antibodies that block Plasmodium vivax transmission.
Article Snippet: Culture supernatants were collected, and cells were lysed in 1× Radioimmunoprecipitation (RIPA) buffer (Millipore 20–188) containing Protease Inhibitor Cocktail III (1:500, Millipore 539,134) and 1 mM Phenylmethanesulfonyl fluoride (PMSF) in isopropanol (Sigma-Aldrich 78,830). .. Proteins were separated SDS-PAGE under reducing condition on 4–20% Mini-PROTEAN TGX Precast Protein Gels (BioRad, Hercules, CA, USA) along with recombinant Pvs25 protein (10 ng) produced by the wheat germ cell-free system (WGCFS) as a positive control [16] and transferred to polyvinylidene fluoride (PVDF) membrane using a Trans-Blot Turbo Transfer System (BioRad) at 1.3A, 25 V for 7 min. Membranes were blocked with BlockPRO Protein-free Blocking Buffer (Energenesis Biomedical, Taipei, Taiwan) for 3 min and incubated with pooled antisera from mice immunized with recombinant Pvs25 (1:20,000) and a mouse anti-β-actin antibody (1:10,000; Abcam ab8226, Cambridge, UK) in 5% (w/v) skim milk in Tris-buffered saline containing Tween 20 (TBST). .. After three washes with TBST, the membranes were incubated with horseradish peroxidase (HRP)-conjugated anti-mouse IgG secondary antibody (1:10,000; Abcam ab6789).
Produced:Article Title: Circular RNA vaccines encoding Pvs25 induce antibodies that block Plasmodium vivax transmission.
Article Snippet: Culture supernatants were collected, and cells were lysed in 1× Radioimmunoprecipitation (RIPA) buffer (Millipore 20–188) containing Protease Inhibitor Cocktail III (1:500, Millipore 539,134) and 1 mM Phenylmethanesulfonyl fluoride (PMSF) in isopropanol (Sigma-Aldrich 78,830). .. Proteins were separated SDS-PAGE under reducing condition on 4–20% Mini-PROTEAN TGX Precast Protein Gels (BioRad, Hercules, CA, USA) along with recombinant Pvs25 protein (10 ng) produced by the wheat germ cell-free system (WGCFS) as a positive control [16] and transferred to polyvinylidene fluoride (PVDF) membrane using a Trans-Blot Turbo Transfer System (BioRad) at 1.3A, 25 V for 7 min. Membranes were blocked with BlockPRO Protein-free Blocking Buffer (Energenesis Biomedical, Taipei, Taiwan) for 3 min and incubated with pooled antisera from mice immunized with recombinant Pvs25 (1:20,000) and a mouse anti-β-actin antibody (1:10,000; Abcam ab8226, Cambridge, UK) in 5% (w/v) skim milk in Tris-buffered saline containing Tween 20 (TBST). .. After three washes with TBST, the membranes were incubated with horseradish peroxidase (HRP)-conjugated anti-mouse IgG secondary antibody (1:10,000; Abcam ab6789).
Positive Control:Article Title: Circular RNA vaccines encoding Pvs25 induce antibodies that block Plasmodium vivax transmission.
Article Snippet: Culture supernatants were collected, and cells were lysed in 1× Radioimmunoprecipitation (RIPA) buffer (Millipore 20–188) containing Protease Inhibitor Cocktail III (1:500, Millipore 539,134) and 1 mM Phenylmethanesulfonyl fluoride (PMSF) in isopropanol (Sigma-Aldrich 78,830). .. Proteins were separated SDS-PAGE under reducing condition on 4–20% Mini-PROTEAN TGX Precast Protein Gels (BioRad, Hercules, CA, USA) along with recombinant Pvs25 protein (10 ng) produced by the wheat germ cell-free system (WGCFS) as a positive control [16] and transferred to polyvinylidene fluoride (PVDF) membrane using a Trans-Blot Turbo Transfer System (BioRad) at 1.3A, 25 V for 7 min. Membranes were blocked with BlockPRO Protein-free Blocking Buffer (Energenesis Biomedical, Taipei, Taiwan) for 3 min and incubated with pooled antisera from mice immunized with recombinant Pvs25 (1:20,000) and a mouse anti-β-actin antibody (1:10,000; Abcam ab8226, Cambridge, UK) in 5% (w/v) skim milk in Tris-buffered saline containing Tween 20 (TBST). .. After three washes with TBST, the membranes were incubated with horseradish peroxidase (HRP)-conjugated anti-mouse IgG secondary antibody (1:10,000; Abcam ab6789).
Blocking Assay:Article Title: Circular RNA vaccines encoding Pvs25 induce antibodies that block Plasmodium vivax transmission.
Article Snippet: Culture supernatants were collected, and cells were lysed in 1× Radioimmunoprecipitation (RIPA) buffer (Millipore 20–188) containing Protease Inhibitor Cocktail III (1:500, Millipore 539,134) and 1 mM Phenylmethanesulfonyl fluoride (PMSF) in isopropanol (Sigma-Aldrich 78,830). .. Proteins were separated SDS-PAGE under reducing condition on 4–20% Mini-PROTEAN TGX Precast Protein Gels (BioRad, Hercules, CA, USA) along with recombinant Pvs25 protein (10 ng) produced by the wheat germ cell-free system (WGCFS) as a positive control [16] and transferred to polyvinylidene fluoride (PVDF) membrane using a Trans-Blot Turbo Transfer System (BioRad) at 1.3A, 25 V for 7 min. Membranes were blocked with BlockPRO Protein-free Blocking Buffer (Energenesis Biomedical, Taipei, Taiwan) for 3 min and incubated with pooled antisera from mice immunized with recombinant Pvs25 (1:20,000) and a mouse anti-β-actin antibody (1:10,000; Abcam ab8226, Cambridge, UK) in 5% (w/v) skim milk in Tris-buffered saline containing Tween 20 (TBST). .. After three washes with TBST, the membranes were incubated with horseradish peroxidase (HRP)-conjugated anti-mouse IgG secondary antibody (1:10,000; Abcam ab6789).
Incubation:Article Title: Circular RNA vaccines encoding Pvs25 induce antibodies that block Plasmodium vivax transmission.
Article Snippet: Culture supernatants were collected, and cells were lysed in 1× Radioimmunoprecipitation (RIPA) buffer (Millipore 20–188) containing Protease Inhibitor Cocktail III (1:500, Millipore 539,134) and 1 mM Phenylmethanesulfonyl fluoride (PMSF) in isopropanol (Sigma-Aldrich 78,830). .. Proteins were separated SDS-PAGE under reducing condition on 4–20% Mini-PROTEAN TGX Precast Protein Gels (BioRad, Hercules, CA, USA) along with recombinant Pvs25 protein (10 ng) produced by the wheat germ cell-free system (WGCFS) as a positive control [16] and transferred to polyvinylidene fluoride (PVDF) membrane using a Trans-Blot Turbo Transfer System (BioRad) at 1.3A, 25 V for 7 min. Membranes were blocked with BlockPRO Protein-free Blocking Buffer (Energenesis Biomedical, Taipei, Taiwan) for 3 min and incubated with pooled antisera from mice immunized with recombinant Pvs25 (1:20,000) and a mouse anti-β-actin antibody (1:10,000; Abcam ab8226, Cambridge, UK) in 5% (w/v) skim milk in Tris-buffered saline containing Tween 20 (TBST). .. After three washes with TBST, the membranes were incubated with horseradish peroxidase (HRP)-conjugated anti-mouse IgG secondary antibody (1:10,000; Abcam ab6789).
Saline:Article Title: Circular RNA vaccines encoding Pvs25 induce antibodies that block Plasmodium vivax transmission.
Article Snippet: Culture supernatants were collected, and cells were lysed in 1× Radioimmunoprecipitation (RIPA) buffer (Millipore 20–188) containing Protease Inhibitor Cocktail III (1:500, Millipore 539,134) and 1 mM Phenylmethanesulfonyl fluoride (PMSF) in isopropanol (Sigma-Aldrich 78,830). .. Proteins were separated SDS-PAGE under reducing condition on 4–20% Mini-PROTEAN TGX Precast Protein Gels (BioRad, Hercules, CA, USA) along with recombinant Pvs25 protein (10 ng) produced by the wheat germ cell-free system (WGCFS) as a positive control [16] and transferred to polyvinylidene fluoride (PVDF) membrane using a Trans-Blot Turbo Transfer System (BioRad) at 1.3A, 25 V for 7 min. Membranes were blocked with BlockPRO Protein-free Blocking Buffer (Energenesis Biomedical, Taipei, Taiwan) for 3 min and incubated with pooled antisera from mice immunized with recombinant Pvs25 (1:20,000) and a mouse anti-β-actin antibody (1:10,000; Abcam ab8226, Cambridge, UK) in 5% (w/v) skim milk in Tris-buffered saline containing Tween 20 (TBST). .. After three washes with TBST, the membranes were incubated with horseradish peroxidase (HRP)-conjugated anti-mouse IgG secondary antibody (1:10,000; Abcam ab6789).
Staining:Article Title: Anti-mycobacterial activity of phage-encoded lysterases.
Article Snippet: .. Heat-denatured samples were collected, loaded into 4%–20% Mini-PROTEAN TGX Precast Protein Gels (#4561094, BioRad), and electrophoresed at 250 V for 15 min. Gels were subsequently stained with Imperial Protein Stain (#24615, ThermoFisher) and destained with deionized water, according to the manufacturer’s protocol. .. The stained protein bands were visualized under a GelDoc system (Universal Hood II, Bio-Rad) and Image Lab software (v 3.0 build 11).
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