ethylenediaminetetraacetic acid disodium salt  (Millipore)


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    Name:
    Ethylenediaminetetraacetic acid disodium salt solution
    Description:
    Ethylenediamine tetraacetate EDTA is a calcium ion chelator that has a low molecular mass of 292 24 Da
    Catalog Number:
    2854
    Price:
    None
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    Structured Review

    Millipore ethylenediaminetetraacetic acid disodium salt
    Ethylenediaminetetraacetic acid disodium salt solution
    Ethylenediamine tetraacetate EDTA is a calcium ion chelator that has a low molecular mass of 292 24 Da
    https://www.bioz.com/result/ethylenediaminetetraacetic acid disodium salt/product/Millipore
    Average 99 stars, based on 3 article reviews
    Price from $9.99 to $1999.99
    ethylenediaminetetraacetic acid disodium salt - by Bioz Stars, 2020-03
    99/100 stars

    Images

    1) Product Images from "Measuring Thiols in Single Cultivar South African Red Wines Using 4,4-Dithiodipyridine (DTDP) Derivatization and Ultraperformance Convergence Chromatography-Tandem Mass Spectrometry"

    Article Title: Measuring Thiols in Single Cultivar South African Red Wines Using 4,4-Dithiodipyridine (DTDP) Derivatization and Ultraperformance Convergence Chromatography-Tandem Mass Spectrometry

    Journal: Foods

    doi: 10.3390/foods7090138

    Sample preparation and instrumental analysis of varietal thiols in white and red wine through 4,4-dithiodipyridine (DTDP) derivatization. EDTA, ethylenediaminetetraacetic acid disodium salt; SPE, solid phase extraction; RT, room temperature; UPC 2 , ultraperformance convergence chromatography; MS/MS, tandem mass spectrometry; 3-MH: 3-mercaptohexanol; 3-MHA: 3-mercaptohexyl acetate; 4-MMP: 4-mercapto-4-methylpentane-2-one; FMT; furanmethanethiol; the DTDP appendix designates the respective derivative.
    Figure Legend Snippet: Sample preparation and instrumental analysis of varietal thiols in white and red wine through 4,4-dithiodipyridine (DTDP) derivatization. EDTA, ethylenediaminetetraacetic acid disodium salt; SPE, solid phase extraction; RT, room temperature; UPC 2 , ultraperformance convergence chromatography; MS/MS, tandem mass spectrometry; 3-MH: 3-mercaptohexanol; 3-MHA: 3-mercaptohexyl acetate; 4-MMP: 4-mercapto-4-methylpentane-2-one; FMT; furanmethanethiol; the DTDP appendix designates the respective derivative.

    Techniques Used: Sample Prep, Chromatography, Mass Spectrometry

    Related Articles

    High Performance Liquid Chromatography:

    Article Title: Ultrasound-Assisted Extraction of Total Flavonoids from Pteris cretica L.: Process Optimization, HPLC Analysis, and Evaluation of Antioxidant Activity
    Article Snippet: HPLC grade trifluoroacetic acid (TFA) and methanol were obtained from Aladdin Reagent Co., Ltd. (Shanghai, China). .. Ascrobic acid, 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), ethylenediaminetetraacetic acid disodium salt (EDTA-2Na), sodium nitroprusside (SNP), potassium persulfate (K2 S2 O8 ), and trolox were obtained from Sigma-Aldrich (St. Louis, MO, USA).

    Article Title: A comparative method for protein extraction and 2-D gel electrophoresis from different tissues of Cajanus cajan
    Article Snippet: Reagents The chemicals and the reagents used in the experiments were HPLC grade and procured from Sigma-Aldrich (≥99.93%). .. 2-Mercaptoethanol (2-ME, Sigma-Aldrich, cat. no. M6250), 3-[(3-Cholamidopropyl) dimethylammonio]-1-propanesulfonate (CHAPS, Sigma-Aldrich, cat. no. C3023), Acetone (Sigma-Aldrich, cat. no. 650501), Acetic acid (Sigma-Aldrich, cat. no. V800018), Agarose (Sigma-Aldrich, cat. no. A9539), Ammonium acetate (Sigma-Aldrich, cat. no. A7262), Bradford reagent (Sigma-Aldrich, cat. no. B6916), Bromophenol blue (BPB, Sigma-Aldrich, cat. no. B0126), Bovine serum albumin (BSA, Sigma-Aldrich, cat. no. 05470), Coomassie brilliant blue (CBB, Bioworld, cat. no. 742083), Dithiothreitol (DTT, Sigma-Aldrich, cat. no. D9779), Ethylene diamine tetra-acetic acid, 0.5 M, pH 8.0 (EDTA, Sigma-Aldrich, cat. no. E7889), Glycerol (Sigma-Aldrich, cat. no. G5516), Glycine (Sigma-Aldrich, cat. no. 50046), Hydrogen chloride (HCl, Sigma-Aldrich, cat. no. H1758), ImmobilineTM Dry strip, pH 3-10, 13 cm (GE Healthcare, UK, cat. no. 17-6001-14), IPG buffer, pH 3-10 NL (GE Healthcare, UK, cat. no. 17-6000-88), Iodoacetamide (IAA, Sigma-Aldrich, cat. no. I3750), Methanol (Sigma-Aldrich, cat. no. 322415), Phenyl methane sulfonyl fluoride (PMSF, Sigma-Aldrich, cat. no. P7626), Potassium chloride (KCl, Sigma-Aldrich, cat. no. P9541), Phosphate buffer, 1M, pH 7.5 (Sigma-Aldrich, cat. no P3619), Complete EDTA free Protease inhibitor cocktail (Roche, cat. no. REF 11 875 580 001), Sodium dodecyl sulfate (SDS, Sigma-Aldrich, cat. no. L4390), Sucrose (Sigma-Aldrich, cat. no. S9378), Thiourea (Sigma-Aldrich, cat. no. T8656), Trichloroacetic acid (TCA; Sigma-Aldrich, cat. no. 522082), Tris (Trizma base, Sigma-Aldrich, cat. no. T1503), Tris-buffered phenol solution (stored at 4°C, pH 7.8–8.0; Sigma-Aldrich, cat. no. P 4557), Tris-HCl, 1.5 M, pH 8.8 (Amresco, cat. no. M195), Urea (Sigma-Aldrich, cat. no. U6504).

    MTT Assay:

    Article Title: Benzylidene derivatives of andrographolide inhibit growth of breast and colon cancer cells in vitro by inducing G1 arrest and apoptosis
    Article Snippet: .. Acridine orange, ammonium persulphate, bovine serum albumin, bromophenol blue, DMSO, N , N , N′ , N′ -tetramethylethylenediamine (TEMED), polyoxyethylene sorbitan monolaurate (Tween 20), PI, MTT, ethyleneglycol-bis-(β-amino ethylether)- N , N , N′ , N′ -tetraacetic acid (EGTA), glycerol, leupeptin, 2-mercaptoethanol, N , N′ -methylene-bis-acrylamide, sodium dodecyl sulphate (SDS), SRB, Tris-base, Triton X-100 and trypsin-ethylenediaminetetra-acetic acid disodium salt (EDTA) were purchased from Sigma (Poole, UK). .. Glycine, sodium chloride, sodium hydroxide and hydrochloric acid were supplied by Fisher Scientific (Loughborough, UK).

    Liquid Chromatography with Mass Spectroscopy:

    Article Title: Identification of multiple serine to asparagine sequence variation sites in an intended copy product of LUCENTIS® by mass spectrometry
    Article Snippet: .. Reagents Lys-C (MS grade, 125–05061) was purchased from Wako, 8 M guanidine hydrochloride (24115) was purchased from Pierce, ethylenediaminetetraacetic acid (EDTA) disodium salt, dehydrate (E5134–250G) was purchased from Sigma, 1 M Tris-hydrochloride pH8 solution (Ultrapure, 15568–025) was purchased from Gibco, dithiothreitol (43815–1G) was purchased from Sigma, iodoacetamide (Bioultra, I1149–5G) was purchased from Sigma, acetonitrile (ACN; LC-MS grade, 9821) was purchased from Biosolve, water (LC-MS grade, 9823–02) was purchased from J.T.Baker, isopropyl alcohol (IPA; LC-MS grade, 34965) from Fluka, and TFA ampoules (MS grade, 28904) were purchased from Pierce. .. Potency assays Two different methods were used for potency determinations, a target binding assay and a cell-based functional assay.

    Modification:

    Article Title: Release Characteristics and Osteogenic Activity of Recombinant Human Bone Morphogenetic Protein-2 grafted to Novel Self-Assembled Poly(lactide-co-glycolide fumarate) Nanoparticles
    Article Snippet: Ethylenediaminetetraacetic acid disodium salt (EDTA), penicillin, streptomycin, Alizarin red, and paraformaldehyde were purchased from Sigma (St. Louis, MO). .. Dulbecco’s phosphate-buffered saline (PBS) and Dulbecco’s Modified Eagle’s Medium (DMEM; 4.5 g/L glucose with L-glutamine and without sodium pyruvate) were obtained from GIBCO BRL (Grand Island, NY).

    Article Title: Erythrocyte Membrane Modified Janus Polymeric Motors for Thrombus Therapy
    Article Snippet: Materials Poly(ethylene imine) solution (M w = 750 000 g/mol), chitosan (medium molecular weight), heparin sodium salt (from porcine intestinal mucosa), phenylmethanesulfonyl fluoride, sodium bicarbonate, ethylenediaminetetraacetic acid disodium salt dehydrate, rhodamine B isothiocyanate, sodium chloride, ammonium fluoride (NH4 F), and fibrinogen from human plasma were obtained from Sigma-Aldrich. .. Protein BCA protein assay kit, Dulbecco’s modified Eagle’s medium (DMEM, high glucose), PBS (1 ×, pH 7.4), cell membrane marker (Wheat germ agglutinin, Alexa Fluor 488 conjugate), thrombin, and Alexa Fluor 488-conjugated fibrinogen were purchased from ThermoFisher Scientific.

    Sulforhodamine B Assay:

    Article Title: Benzylidene derivatives of andrographolide inhibit growth of breast and colon cancer cells in vitro by inducing G1 arrest and apoptosis
    Article Snippet: .. Acridine orange, ammonium persulphate, bovine serum albumin, bromophenol blue, DMSO, N , N , N′ , N′ -tetramethylethylenediamine (TEMED), polyoxyethylene sorbitan monolaurate (Tween 20), PI, MTT, ethyleneglycol-bis-(β-amino ethylether)- N , N , N′ , N′ -tetraacetic acid (EGTA), glycerol, leupeptin, 2-mercaptoethanol, N , N′ -methylene-bis-acrylamide, sodium dodecyl sulphate (SDS), SRB, Tris-base, Triton X-100 and trypsin-ethylenediaminetetra-acetic acid disodium salt (EDTA) were purchased from Sigma (Poole, UK). .. Glycine, sodium chloride, sodium hydroxide and hydrochloric acid were supplied by Fisher Scientific (Loughborough, UK).

    Multiple Displacement Amplification:

    Article Title: Optimum 3D Matrix Stiffness for Maintenance of Cancer Stem Cells Is Dependent on Tissue Origin of Cancer Cells
    Article Snippet: Piperidine, tetrahydrofuran (THF), trimethylsilane (TMS), triethylamine (TEA), acrylic acid, dimethylsulfoxide (DMSO), and ethylenediaminetetraacetic acid disodium salt (EDTA) were from Sigma-Aldrich. .. MCF7 (HTB-22) breast adenocarcinoma, MDA-MB-231 (HTB-26, hereafter denoted by MDA231) breast adenocarcinoma, HCT116 (CCL-247) colorectal carcinoma, AGS (CRL-1739) gastric adenocarcinoma, U2OS (HTB-96) osteosarcoma cell lines, and MCF10A (CRL-10317) non-tumorigenic epithelial cells were purchased from ATCC (Manassas, VA).

    Isolation:

    Article Title: Influence of Solvent Polarity and DNA-Binding on Spectral Properties of Quaternary Benzo[c]phenanthridine Alkaloids
    Article Snippet: Quaternary benzo[c ]phenanthridine alkaloids and chemicals All studied alkaloids were isolated from plant material as published earlier [ ] and were obtained as chloride salts of at least 93% purity. .. Trizma base, ethylenediaminetetraacetic acid disodium salt (EDTA), quinine, colloidal silica (Ludox), calf thymus DNA (ctDNA), and salmon testes DNA were purchased from Sigma-Aldrich, USA.

    Synthesized:

    Article Title: Benzylidene derivatives of andrographolide inhibit growth of breast and colon cancer cells in vitro by inducing G1 arrest and apoptosis
    Article Snippet: All the andrographolide analogues were synthesized in our laboratory ( ). .. Acridine orange, ammonium persulphate, bovine serum albumin, bromophenol blue, DMSO, N , N , N′ , N′ -tetramethylethylenediamine (TEMED), polyoxyethylene sorbitan monolaurate (Tween 20), PI, MTT, ethyleneglycol-bis-(β-amino ethylether)- N , N , N′ , N′ -tetraacetic acid (EGTA), glycerol, leupeptin, 2-mercaptoethanol, N , N′ -methylene-bis-acrylamide, sodium dodecyl sulphate (SDS), SRB, Tris-base, Triton X-100 and trypsin-ethylenediaminetetra-acetic acid disodium salt (EDTA) were purchased from Sigma (Poole, UK).

    Distillation:

    Article Title: Release Characteristics and Osteogenic Activity of Recombinant Human Bone Morphogenetic Protein-2 grafted to Novel Self-Assembled Poly(lactide-co-glycolide fumarate) Nanoparticles
    Article Snippet: DCM was dried by distillation over calcium hydride. .. Ethylenediaminetetraacetic acid disodium salt (EDTA), penicillin, streptomycin, Alizarin red, and paraformaldehyde were purchased from Sigma (St. Louis, MO).

    Purification:

    Article Title: Assessment of BOLD and GenBank – Their accuracy and reliability for the identification of biological materials
    Article Snippet: The washed pellet was subsequently dried using a 55°C hot plate (~5 min) and re-suspended in 50 μL of TE Buffer (10mM Trizma HCl (Sigma-Aldrich [T3038]), 1mM EDTA (Sigma-Aldrich [E7889])). .. Extracts were purified with Agencourt AMPure XP beads (Beckman Coulter [A63880], Brea, CA, USA) per manufacturer’s recommendations for genomic DNA.

    Concentration Assay:

    Article Title: Influence of Solvent Polarity and DNA-Binding on Spectral Properties of Quaternary Benzo[c]phenanthridine Alkaloids
    Article Snippet: Trizma base, ethylenediaminetetraacetic acid disodium salt (EDTA), quinine, colloidal silica (Ludox), calf thymus DNA (ctDNA), and salmon testes DNA were purchased from Sigma-Aldrich, USA. .. The concentration of DNA was determined spectrophotometrically using the relationship that 1 absorbance unit at 260 nm corresponds to 50 μg ml-1 (0.075 mM in base-pairs (bp)) of double-stranded DNA.

    Incubation:

    Article Title: Assessment of BOLD and GenBank – Their accuracy and reliability for the identification of biological materials
    Article Snippet: Following incubation, 500 μL of Phenol:Chloroform:Isoamylalcohol (25:24:1; Sigma-Aldrich [AM9732]) was added, mixed well and centrifuged for 10 min at 12,000 x g . .. The washed pellet was subsequently dried using a 55°C hot plate (~5 min) and re-suspended in 50 μL of TE Buffer (10mM Trizma HCl (Sigma-Aldrich [T3038]), 1mM EDTA (Sigma-Aldrich [E7889])).

    other:

    Article Title: Analysis of the Link between Enzymatic Activity and Oligomeric State in AhpC, a Bacterial Peroxiredoxin of the Link between Enzymatic Activity and Oligomeric State in AhpC, a Bacterial Peroxiredoxin
    Article Snippet: Sodium dodecyl sulfate (SDS), ultrapure glycine, ethylenediaminetetraacetic acid (EDTA) disodium salt, dithiothreitol (DTT), ammonium sulfate, Tris base, and other buffer reagents were purchased from Research Organics (Cleveland, OH).

    Article Title: Measuring Thiols in Single Cultivar South African Red Wines Using 4,4-Dithiodipyridine (DTDP) Derivatization and Ultraperformance Convergence Chromatography-Tandem Mass Spectrometry
    Article Snippet: All reagents—3-MH, 6-mercapto-1-hexanol (6-MH), 3-MHA, 4-MMP, FMT, 98% DTDP, ethylenediaminetetraacetic acid disodium salt (EDTA–Na2 ), methanol, 96% ethanol, sodium hydroxide (NaOH), tartaric acid, anhydrous acetaldehyde ≥98%, and 37% hydrochloric acid (HCl)—were purchased from Sigma-Aldrich (Louisville, MO, USA) and the solid phase extraction (SPE) cartridges (Supelclean ENVI-18 SPE) from Supelco (Bellefonte, PA, USA ).

    Article Title: Biochemical Properties of Polyphenol Oxidases from Ready-to-Eat Lentil (Lens culinaris Medik.) Sprouts and Factors Affecting Their Activities: A Search for Potent Tools Limiting Enzymatic Browning
    Article Snippet: Chemicals Catechol, Diethylaminoethyl–Sepharose (DEAE–S), tris(hydroxymethyl)aminomethane (TRIS), ethylenediaminetetraacetic acid sodium salt (EDTA), 4-methylcatechol, gallic acid, caffeic acid, l -cysteine, ascorbic acid, and dl -dithiothreitol were obtained from Sigma-Aldrich (Poznań, Poland).

    Article Title: Analysis of Tetracyclines in Medicated Feed for Food Animal Production by HPLC-MS/MS
    Article Snippet: Disodium hydrogen phosphate dehydrate, anhydrous citric acid, ethylenediaminetetraacetic acid disodium salt (EDTA), trichloroacetic acid (TCA) and formic acid (purity > 99% by analysis) were purchased from Sigma-Aldrich (St Louis, MO, USA).

    Stripping Membranes:

    Article Title: A comparative method for protein extraction and 2-D gel electrophoresis from different tissues of Cajanus cajan
    Article Snippet: .. 2-Mercaptoethanol (2-ME, Sigma-Aldrich, cat. no. M6250), 3-[(3-Cholamidopropyl) dimethylammonio]-1-propanesulfonate (CHAPS, Sigma-Aldrich, cat. no. C3023), Acetone (Sigma-Aldrich, cat. no. 650501), Acetic acid (Sigma-Aldrich, cat. no. V800018), Agarose (Sigma-Aldrich, cat. no. A9539), Ammonium acetate (Sigma-Aldrich, cat. no. A7262), Bradford reagent (Sigma-Aldrich, cat. no. B6916), Bromophenol blue (BPB, Sigma-Aldrich, cat. no. B0126), Bovine serum albumin (BSA, Sigma-Aldrich, cat. no. 05470), Coomassie brilliant blue (CBB, Bioworld, cat. no. 742083), Dithiothreitol (DTT, Sigma-Aldrich, cat. no. D9779), Ethylene diamine tetra-acetic acid, 0.5 M, pH 8.0 (EDTA, Sigma-Aldrich, cat. no. E7889), Glycerol (Sigma-Aldrich, cat. no. G5516), Glycine (Sigma-Aldrich, cat. no. 50046), Hydrogen chloride (HCl, Sigma-Aldrich, cat. no. H1758), ImmobilineTM Dry strip, pH 3-10, 13 cm (GE Healthcare, UK, cat. no. 17-6001-14), IPG buffer, pH 3-10 NL (GE Healthcare, UK, cat. no. 17-6000-88), Iodoacetamide (IAA, Sigma-Aldrich, cat. no. I3750), Methanol (Sigma-Aldrich, cat. no. 322415), Phenyl methane sulfonyl fluoride (PMSF, Sigma-Aldrich, cat. no. P7626), Potassium chloride (KCl, Sigma-Aldrich, cat. no. P9541), Phosphate buffer, 1M, pH 7.5 (Sigma-Aldrich, cat. no P3619), Complete EDTA free Protease inhibitor cocktail (Roche, cat. no. REF 11 875 580 001), Sodium dodecyl sulfate (SDS, Sigma-Aldrich, cat. no. L4390), Sucrose (Sigma-Aldrich, cat. no. S9378), Thiourea (Sigma-Aldrich, cat. no. T8656), Trichloroacetic acid (TCA; Sigma-Aldrich, cat. no. 522082), Tris (Trizma base, Sigma-Aldrich, cat. no. T1503), Tris-buffered phenol solution (stored at 4°C, pH 7.8–8.0; Sigma-Aldrich, cat. no. P 4557), Tris-HCl, 1.5 M, pH 8.8 (Amresco, cat. no. M195), Urea (Sigma-Aldrich, cat. no. U6504). ..

    Mass Spectrometry:

    Article Title: Identification of multiple serine to asparagine sequence variation sites in an intended copy product of LUCENTIS® by mass spectrometry
    Article Snippet: .. Reagents Lys-C (MS grade, 125–05061) was purchased from Wako, 8 M guanidine hydrochloride (24115) was purchased from Pierce, ethylenediaminetetraacetic acid (EDTA) disodium salt, dehydrate (E5134–250G) was purchased from Sigma, 1 M Tris-hydrochloride pH8 solution (Ultrapure, 15568–025) was purchased from Gibco, dithiothreitol (43815–1G) was purchased from Sigma, iodoacetamide (Bioultra, I1149–5G) was purchased from Sigma, acetonitrile (ACN; LC-MS grade, 9821) was purchased from Biosolve, water (LC-MS grade, 9823–02) was purchased from J.T.Baker, isopropyl alcohol (IPA; LC-MS grade, 34965) from Fluka, and TFA ampoules (MS grade, 28904) were purchased from Pierce. .. Potency assays Two different methods were used for potency determinations, a target binding assay and a cell-based functional assay.

    Protease Inhibitor:

    Article Title: A comparative method for protein extraction and 2-D gel electrophoresis from different tissues of Cajanus cajan
    Article Snippet: .. 2-Mercaptoethanol (2-ME, Sigma-Aldrich, cat. no. M6250), 3-[(3-Cholamidopropyl) dimethylammonio]-1-propanesulfonate (CHAPS, Sigma-Aldrich, cat. no. C3023), Acetone (Sigma-Aldrich, cat. no. 650501), Acetic acid (Sigma-Aldrich, cat. no. V800018), Agarose (Sigma-Aldrich, cat. no. A9539), Ammonium acetate (Sigma-Aldrich, cat. no. A7262), Bradford reagent (Sigma-Aldrich, cat. no. B6916), Bromophenol blue (BPB, Sigma-Aldrich, cat. no. B0126), Bovine serum albumin (BSA, Sigma-Aldrich, cat. no. 05470), Coomassie brilliant blue (CBB, Bioworld, cat. no. 742083), Dithiothreitol (DTT, Sigma-Aldrich, cat. no. D9779), Ethylene diamine tetra-acetic acid, 0.5 M, pH 8.0 (EDTA, Sigma-Aldrich, cat. no. E7889), Glycerol (Sigma-Aldrich, cat. no. G5516), Glycine (Sigma-Aldrich, cat. no. 50046), Hydrogen chloride (HCl, Sigma-Aldrich, cat. no. H1758), ImmobilineTM Dry strip, pH 3-10, 13 cm (GE Healthcare, UK, cat. no. 17-6001-14), IPG buffer, pH 3-10 NL (GE Healthcare, UK, cat. no. 17-6000-88), Iodoacetamide (IAA, Sigma-Aldrich, cat. no. I3750), Methanol (Sigma-Aldrich, cat. no. 322415), Phenyl methane sulfonyl fluoride (PMSF, Sigma-Aldrich, cat. no. P7626), Potassium chloride (KCl, Sigma-Aldrich, cat. no. P9541), Phosphate buffer, 1M, pH 7.5 (Sigma-Aldrich, cat. no P3619), Complete EDTA free Protease inhibitor cocktail (Roche, cat. no. REF 11 875 580 001), Sodium dodecyl sulfate (SDS, Sigma-Aldrich, cat. no. L4390), Sucrose (Sigma-Aldrich, cat. no. S9378), Thiourea (Sigma-Aldrich, cat. no. T8656), Trichloroacetic acid (TCA; Sigma-Aldrich, cat. no. 522082), Tris (Trizma base, Sigma-Aldrich, cat. no. T1503), Tris-buffered phenol solution (stored at 4°C, pH 7.8–8.0; Sigma-Aldrich, cat. no. P 4557), Tris-HCl, 1.5 M, pH 8.8 (Amresco, cat. no. M195), Urea (Sigma-Aldrich, cat. no. U6504). ..

    Marker:

    Article Title: Erythrocyte Membrane Modified Janus Polymeric Motors for Thrombus Therapy
    Article Snippet: Materials Poly(ethylene imine) solution (M w = 750 000 g/mol), chitosan (medium molecular weight), heparin sodium salt (from porcine intestinal mucosa), phenylmethanesulfonyl fluoride, sodium bicarbonate, ethylenediaminetetraacetic acid disodium salt dehydrate, rhodamine B isothiocyanate, sodium chloride, ammonium fluoride (NH4 F), and fibrinogen from human plasma were obtained from Sigma-Aldrich. .. Protein BCA protein assay kit, Dulbecco’s modified Eagle’s medium (DMEM, high glucose), PBS (1 ×, pH 7.4), cell membrane marker (Wheat germ agglutinin, Alexa Fluor 488 conjugate), thrombin, and Alexa Fluor 488-conjugated fibrinogen were purchased from ThermoFisher Scientific.

    Staining:

    Article Title: Erythrocyte Membrane Modified Janus Polymeric Motors for Thrombus Therapy
    Article Snippet: Materials Poly(ethylene imine) solution (M w = 750 000 g/mol), chitosan (medium molecular weight), heparin sodium salt (from porcine intestinal mucosa), phenylmethanesulfonyl fluoride, sodium bicarbonate, ethylenediaminetetraacetic acid disodium salt dehydrate, rhodamine B isothiocyanate, sodium chloride, ammonium fluoride (NH4 F), and fibrinogen from human plasma were obtained from Sigma-Aldrich. .. Mini-PROTEAN@ TGX stain-free gels, 4 × Laemmli protein sample buffer, and Precision Plus Protein All Blue Standards were obtained from Bio-Rad.

    BIA-KA:

    Article Title: Erythrocyte Membrane Modified Janus Polymeric Motors for Thrombus Therapy
    Article Snippet: Materials Poly(ethylene imine) solution (M w = 750 000 g/mol), chitosan (medium molecular weight), heparin sodium salt (from porcine intestinal mucosa), phenylmethanesulfonyl fluoride, sodium bicarbonate, ethylenediaminetetraacetic acid disodium salt dehydrate, rhodamine B isothiocyanate, sodium chloride, ammonium fluoride (NH4 F), and fibrinogen from human plasma were obtained from Sigma-Aldrich. .. Protein BCA protein assay kit, Dulbecco’s modified Eagle’s medium (DMEM, high glucose), PBS (1 ×, pH 7.4), cell membrane marker (Wheat germ agglutinin, Alexa Fluor 488 conjugate), thrombin, and Alexa Fluor 488-conjugated fibrinogen were purchased from ThermoFisher Scientific.

    Indirect Immunoperoxidase Assay:

    Article Title: Identification of multiple serine to asparagine sequence variation sites in an intended copy product of LUCENTIS® by mass spectrometry
    Article Snippet: .. Reagents Lys-C (MS grade, 125–05061) was purchased from Wako, 8 M guanidine hydrochloride (24115) was purchased from Pierce, ethylenediaminetetraacetic acid (EDTA) disodium salt, dehydrate (E5134–250G) was purchased from Sigma, 1 M Tris-hydrochloride pH8 solution (Ultrapure, 15568–025) was purchased from Gibco, dithiothreitol (43815–1G) was purchased from Sigma, iodoacetamide (Bioultra, I1149–5G) was purchased from Sigma, acetonitrile (ACN; LC-MS grade, 9821) was purchased from Biosolve, water (LC-MS grade, 9823–02) was purchased from J.T.Baker, isopropyl alcohol (IPA; LC-MS grade, 34965) from Fluka, and TFA ampoules (MS grade, 28904) were purchased from Pierce. .. Potency assays Two different methods were used for potency determinations, a target binding assay and a cell-based functional assay.

    Molecular Weight:

    Article Title: Optimum 3D Matrix Stiffness for Maintenance of Cancer Stem Cells Is Dependent on Tissue Origin of Cancer Cells
    Article Snippet: Materials Linear polyethylene glycol (PEG) with molecular weight (MW) of 3.4 kDa was purchased from Acros (Pittsburg, PA). .. Piperidine, tetrahydrofuran (THF), trimethylsilane (TMS), triethylamine (TEA), acrylic acid, dimethylsulfoxide (DMSO), and ethylenediaminetetraacetic acid disodium salt (EDTA) were from Sigma-Aldrich.

    Article Title: Release Characteristics and Osteogenic Activity of Recombinant Human Bone Morphogenetic Protein-2 grafted to Novel Self-Assembled Poly(lactide-co-glycolide fumarate) Nanoparticles
    Article Snippet: Spectro/Por dialysis tube (molecular weight cut-off 3.5 kDa) was purchased from Spectrum Laboratories (Rancho Dominquez, CA). .. Ethylenediaminetetraacetic acid disodium salt (EDTA), penicillin, streptomycin, Alizarin red, and paraformaldehyde were purchased from Sigma (St. Louis, MO).

    Article Title: Erythrocyte Membrane Modified Janus Polymeric Motors for Thrombus Therapy
    Article Snippet: .. Materials Poly(ethylene imine) solution (M w = 750 000 g/mol), chitosan (medium molecular weight), heparin sodium salt (from porcine intestinal mucosa), phenylmethanesulfonyl fluoride, sodium bicarbonate, ethylenediaminetetraacetic acid disodium salt dehydrate, rhodamine B isothiocyanate, sodium chloride, ammonium fluoride (NH4 F), and fibrinogen from human plasma were obtained from Sigma-Aldrich. .. Silica spheres with a diameter of 5 μm were obtained from Microparticles GmbH, Germany.

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  • 99
    Millipore nmr buffer
    The m 6 A modification has minor effects on RREIIB structure and dynamics. (A) Secondary structure of RRE2Bm 6A68 , with the residues showing line-broadening with m 6 A, highlighted in red (with Mg 2+ ) and blue (no Mg 2+ ). Comparison of the 1D 1 H <t>NMR</t> spectrum of RREIIB m6A68 with or without Mg 2+ with the methyl peak indicated by arrows. The comparison of 1D imino spectra (B) and 2D [ 1 H, 13 C]-HSQC spectra (C) of RREIIB m6A68 and RREIIB in the presence (red) and absence (blue) of 3 mM Mg 2+ . Resonances exhibiting shifting are indicated using arrows, and those with ambiguous assignments denoted using an asterisk. (D) Normalized resonance intensities in 2D [ 1 H, 13 C]-HSQC spectra of RREIIB m6A68 and RREIIB in the presence (red) and absence (blue) of 3 mM Mg 2+ . A52-C8H8, A52-C2H2 and U56-C6H6 were used as a reference and normalized to 0.1. The sample conditions were 1.2–1.5 mM RREIIB m6A68 or RREIIB in 15 mM sodium phosphate, 25 mM <t>NaCl,</t> 0.1 mM EDTA, pH 6.4 with or without 3 mM MgCl 2 .
    Nmr Buffer, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 106 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/nmr buffer/product/Millipore
    Average 99 stars, based on 106 article reviews
    Price from $9.99 to $1999.99
    nmr buffer - by Bioz Stars, 2020-03
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    99
    Millipore calpain reaction buffer
    Schematic diagram showing the relationship of <t>calpain/NF-κB/inflammation/NVU</t> damage after CCI in mice. Traumatic brain injury induces calcium overload, which, in turn, upregulates calpain. Calpain may downregulate IκB and activate NF-κB. NF-κB induces activation of TNF-α, iNOS, ICAM-1, and MMP-9. These inflammatory substances induce degradation of basal lamina and tight junction proteins, resulting in NVU disruption, leading to brain edema. MDL28170 could reverse those changes. NF-κB: Nuclear factor-κB; NVU: Neurovascular unit; CCI: Controlled cortical impact; IκB: Inhibitory-κB; TNF-α: Tumor necrosis factor-α; iNOS: Inducible nitric oxide synthase; ICAM-1: Intracellular adhesion molecule-1; MMP-9: Matrix metalloproteinase-9.
    Calpain Reaction Buffer, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    79
    Millipore uch buffer
    Insoluble α-syn in diseased brain is ubiquitinated. A: Western blot analysis of biochemically fractionated cingulate cortex from a patient with <t>DLB</t> (DLB-3). Immunoblots were developed with anti-α-syn antibodies LB509 and Syn208 as well as anti-ubiquitin antibody mAb 1510. Twenty μl of LS fraction ( lane 1 ), TX fraction ( lane 2 ), sarkosyl-soluble fraction ( lane 3 ), and SDS-soluble fraction ( lane 4 ) were loaded in separate lanes of 15% SDS-polyacrylamide gels. Note that the SDS-soluble fraction is four times as concentrated as each of the other fractions in that 2.5 ml/g of SDS buffer was used for tissue extraction versus 10 ml/g for each of the other fractions. Arrowhead indicates α-syn monomer (αS) and bracket indicates ubiquitin monomer (Ub). Arrows depict mono-, di-, and tri-ubiquitinated forms of α-syn. B: Western blot analysis (with antibodies Syn208 and mAb 1510) of the SDS-soluble fraction from the cingulate cortex of normal brains (NL-1, NL-2) and DLB brains (DLB-1, DLB-2, and DLB-3). Twenty μl of SDS-soluble fraction was loaded in each lane of a 15% gel. One hundred ng of recombinant human α-syn was loaded in the indicated lanes. C: Western blot analysis of the SDS-soluble fraction from the cingulate cortex of case DLB-1 using various anti-α-syn (LB509, Syn102, Syn211) and anti-ubiquitin (mAb 1510, Conj8) antibodies. Arrows depict mono-, di-, and tri-ubiquitinated forms of α-syn. D: Immunoprecipitation followed by Western blot analysis. α-Syn in the SDS-soluble fraction of the cingulate cortex of DLB-1 was isolated by immunoprecipitation with anti-α-syn antibodies. The sample was analyzed by Western blot analysis using anti-α-syn antibody LB509 and anti-ubiquitin antibody mAb 1510 ( arrows , mono- and di-ubiquitinated forms of α-syn; * and **, possible ubiquitinated forms of α-syn in which ubiquitin moieties may be masking the LB509 epitope; ***, modified form of α-syn, possibly dimerized α-syn). E: Ubiquitinated α-syn from DLB brain can be deubiquitinated by <t>UCH-L1</t> in vitro . SDS-soluble fraction from the cingulate cortex of NL-1 or DLB-1 was untreated or reacted with 50 nmol/L of UCH-L1. The samples were analyzed by Western blot analysis using LB509. R represents a lane loaded with 100 ng of recombinant human α-syn. The mobility of molecular mass markers (kd) is depicted on the left of each panel.
    Uch Buffer, supplied by Millipore, used in various techniques. Bioz Stars score: 79/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore jak2 ip buffer
    Erythropoietin Receptor-Mediated Activation of the <t>JAK2/STAT5,</t> RAS/MAPK, and PI3K/AKT Signaling Pathways is Attenuated by BCR-ABL Kinase Activity in K562 Cells A. Normalized hEPO-mediated (10′ stimulation) JAK2 activation in K562 cells treated for 1hr with either 100nM dasatinib or 1μM imatinib. Data represents the average ± SD (n=3). JAK2 activation was monitored by immunoprecipitation and activation loop (Y1007) phosphorylation. JAK2 activation was normalized to the level of phospho-Y1007 observed in the “DMSO” condition for each experimental replicate. B. Upper: Line diagram representation of duration of serum starve, kinase inhibitor treatment, and growth-factor stimulation. Lower: Western immunoblot analysis of whole cell lysates from K562 cells after short-term and prolonged BCR-ABL inhibition (100nM dasatinib: 2hrs, 4hrs, 8hrs, 24hrs; 0.2% DMSO, 24hrs) followed by hEPO stimulation (10′). C. Normalized hEPO-mediated JAK2 activation in K562 cells after short-term (1hr) and prolonged (24hrs) 100nM dasatinib treatment followed hEPO stimulation (10′). Data is representative of triplicate experimental analysis. JAK2 activation and normalization was performed as in (A). D. Normalized RAS-GTP loading in K562 cells after short-term (1hr) and prolonged (24hrs) 100nM dasatinib treatment followed by hEPO stimulation (10′). Data is representative of triplicate experimental analysis. RAS activation was monitored using a RAS-GTP pulldown assay and RAS-GTP levels were normalized to the “DMSO” condition. E. Normalized hEPO-mediated (10′ stimulation) JAK2 activation in K562 cells pretreated for 24hrs with 0.2% DMSO, 100nM dasatinib, 1μM imatinib, 500nM TG101348, dasatinib/TG101348, or imatinib/TG101348. JAK2 activation was monitored and normalized as in (A). F. Upper: Line diagram representation of duration of serum starve, kinase inhibitor treatment, and growth-factor stimulation. Lower: Western immunoblot analysis of whole cell lysates in K562 cells pretreated for 24hrs with 0.2% DMSO, 100nM dasatinib, 1μM imatinib, 500nM TG101348, dasatinib/TG101348, or imatinib/TG101348 followed by hEPO stimulation (10′).
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    Image Search Results


    The m 6 A modification has minor effects on RREIIB structure and dynamics. (A) Secondary structure of RRE2Bm 6A68 , with the residues showing line-broadening with m 6 A, highlighted in red (with Mg 2+ ) and blue (no Mg 2+ ). Comparison of the 1D 1 H NMR spectrum of RREIIB m6A68 with or without Mg 2+ with the methyl peak indicated by arrows. The comparison of 1D imino spectra (B) and 2D [ 1 H, 13 C]-HSQC spectra (C) of RREIIB m6A68 and RREIIB in the presence (red) and absence (blue) of 3 mM Mg 2+ . Resonances exhibiting shifting are indicated using arrows, and those with ambiguous assignments denoted using an asterisk. (D) Normalized resonance intensities in 2D [ 1 H, 13 C]-HSQC spectra of RREIIB m6A68 and RREIIB in the presence (red) and absence (blue) of 3 mM Mg 2+ . A52-C8H8, A52-C2H2 and U56-C6H6 were used as a reference and normalized to 0.1. The sample conditions were 1.2–1.5 mM RREIIB m6A68 or RREIIB in 15 mM sodium phosphate, 25 mM NaCl, 0.1 mM EDTA, pH 6.4 with or without 3 mM MgCl 2 .

    Journal: PLoS ONE

    Article Title: m6A minimally impacts the structure, dynamics, and Rev ARM binding properties of HIV-1 RRE stem IIB

    doi: 10.1371/journal.pone.0224850

    Figure Lengend Snippet: The m 6 A modification has minor effects on RREIIB structure and dynamics. (A) Secondary structure of RRE2Bm 6A68 , with the residues showing line-broadening with m 6 A, highlighted in red (with Mg 2+ ) and blue (no Mg 2+ ). Comparison of the 1D 1 H NMR spectrum of RREIIB m6A68 with or without Mg 2+ with the methyl peak indicated by arrows. The comparison of 1D imino spectra (B) and 2D [ 1 H, 13 C]-HSQC spectra (C) of RREIIB m6A68 and RREIIB in the presence (red) and absence (blue) of 3 mM Mg 2+ . Resonances exhibiting shifting are indicated using arrows, and those with ambiguous assignments denoted using an asterisk. (D) Normalized resonance intensities in 2D [ 1 H, 13 C]-HSQC spectra of RREIIB m6A68 and RREIIB in the presence (red) and absence (blue) of 3 mM Mg 2+ . A52-C8H8, A52-C2H2 and U56-C6H6 were used as a reference and normalized to 0.1. The sample conditions were 1.2–1.5 mM RREIIB m6A68 or RREIIB in 15 mM sodium phosphate, 25 mM NaCl, 0.1 mM EDTA, pH 6.4 with or without 3 mM MgCl 2 .

    Article Snippet: After measuring the concentration, the RNA samples were buffer-exchanged into NMR buffer (15 mM sodium phosphate, 25 mM NaCl, 0.1 mM EDTA, with or without 3 mM MgCl2 at pH = 6.4) three times using 3kDa Amicon Ultra centrifugal filters (EMD Millipore).

    Techniques: Modification, Nuclear Magnetic Resonance

    Schematic diagram showing the relationship of calpain/NF-κB/inflammation/NVU damage after CCI in mice. Traumatic brain injury induces calcium overload, which, in turn, upregulates calpain. Calpain may downregulate IκB and activate NF-κB. NF-κB induces activation of TNF-α, iNOS, ICAM-1, and MMP-9. These inflammatory substances induce degradation of basal lamina and tight junction proteins, resulting in NVU disruption, leading to brain edema. MDL28170 could reverse those changes. NF-κB: Nuclear factor-κB; NVU: Neurovascular unit; CCI: Controlled cortical impact; IκB: Inhibitory-κB; TNF-α: Tumor necrosis factor-α; iNOS: Inducible nitric oxide synthase; ICAM-1: Intracellular adhesion molecule-1; MMP-9: Matrix metalloproteinase-9.

    Journal: Chinese Medical Journal

    Article Title: Protective Effects of Calpain Inhibition on Neurovascular Unit Injury through Downregulating Nuclear Factor-κB-related Inflammation during Traumatic Brain Injury in Mice

    doi: 10.4103/0366-6999.198001

    Figure Lengend Snippet: Schematic diagram showing the relationship of calpain/NF-κB/inflammation/NVU damage after CCI in mice. Traumatic brain injury induces calcium overload, which, in turn, upregulates calpain. Calpain may downregulate IκB and activate NF-κB. NF-κB induces activation of TNF-α, iNOS, ICAM-1, and MMP-9. These inflammatory substances induce degradation of basal lamina and tight junction proteins, resulting in NVU disruption, leading to brain edema. MDL28170 could reverse those changes. NF-κB: Nuclear factor-κB; NVU: Neurovascular unit; CCI: Controlled cortical impact; IκB: Inhibitory-κB; TNF-α: Tumor necrosis factor-α; iNOS: Inducible nitric oxide synthase; ICAM-1: Intracellular adhesion molecule-1; MMP-9: Matrix metalloproteinase-9.

    Article Snippet: [ ] In brief, cytosolic and mitochondrial proteins (30 μg) were incubated with calpain reaction buffer (20 mmol/L HEPES, 1 mmol/L EDTA, 50 mmol/L NaCl, and 0.1% (v/v) 2-mercaptoethanol, containing 10 μmol/L calpain I fluorescent substrate [Calbiochem Co., La Jolla, CA, USA], pH 7.6).

    Techniques: Mouse Assay, Activation Assay

    MDL28170 treatment suppresses the calpain activity in the cytosolic and mitochondrial fractions and upregulates the expression of calpastatin in the cytosolic fractions. (a and b) The bar graphs reflect the calpain activity in the cytosolic fractions and mitochondrial fractions at 6 h and 24 h. (c) Representative Western blots of calpastatin and β-actin from each group; (d) the results were quantified and are shown as the mean ± SD. * P

    Journal: Chinese Medical Journal

    Article Title: Protective Effects of Calpain Inhibition on Neurovascular Unit Injury through Downregulating Nuclear Factor-κB-related Inflammation during Traumatic Brain Injury in Mice

    doi: 10.4103/0366-6999.198001

    Figure Lengend Snippet: MDL28170 treatment suppresses the calpain activity in the cytosolic and mitochondrial fractions and upregulates the expression of calpastatin in the cytosolic fractions. (a and b) The bar graphs reflect the calpain activity in the cytosolic fractions and mitochondrial fractions at 6 h and 24 h. (c) Representative Western blots of calpastatin and β-actin from each group; (d) the results were quantified and are shown as the mean ± SD. * P

    Article Snippet: [ ] In brief, cytosolic and mitochondrial proteins (30 μg) were incubated with calpain reaction buffer (20 mmol/L HEPES, 1 mmol/L EDTA, 50 mmol/L NaCl, and 0.1% (v/v) 2-mercaptoethanol, containing 10 μmol/L calpain I fluorescent substrate [Calbiochem Co., La Jolla, CA, USA], pH 7.6).

    Techniques: Activity Assay, Expressing, Western Blot

    Insoluble α-syn in diseased brain is ubiquitinated. A: Western blot analysis of biochemically fractionated cingulate cortex from a patient with DLB (DLB-3). Immunoblots were developed with anti-α-syn antibodies LB509 and Syn208 as well as anti-ubiquitin antibody mAb 1510. Twenty μl of LS fraction ( lane 1 ), TX fraction ( lane 2 ), sarkosyl-soluble fraction ( lane 3 ), and SDS-soluble fraction ( lane 4 ) were loaded in separate lanes of 15% SDS-polyacrylamide gels. Note that the SDS-soluble fraction is four times as concentrated as each of the other fractions in that 2.5 ml/g of SDS buffer was used for tissue extraction versus 10 ml/g for each of the other fractions. Arrowhead indicates α-syn monomer (αS) and bracket indicates ubiquitin monomer (Ub). Arrows depict mono-, di-, and tri-ubiquitinated forms of α-syn. B: Western blot analysis (with antibodies Syn208 and mAb 1510) of the SDS-soluble fraction from the cingulate cortex of normal brains (NL-1, NL-2) and DLB brains (DLB-1, DLB-2, and DLB-3). Twenty μl of SDS-soluble fraction was loaded in each lane of a 15% gel. One hundred ng of recombinant human α-syn was loaded in the indicated lanes. C: Western blot analysis of the SDS-soluble fraction from the cingulate cortex of case DLB-1 using various anti-α-syn (LB509, Syn102, Syn211) and anti-ubiquitin (mAb 1510, Conj8) antibodies. Arrows depict mono-, di-, and tri-ubiquitinated forms of α-syn. D: Immunoprecipitation followed by Western blot analysis. α-Syn in the SDS-soluble fraction of the cingulate cortex of DLB-1 was isolated by immunoprecipitation with anti-α-syn antibodies. The sample was analyzed by Western blot analysis using anti-α-syn antibody LB509 and anti-ubiquitin antibody mAb 1510 ( arrows , mono- and di-ubiquitinated forms of α-syn; * and **, possible ubiquitinated forms of α-syn in which ubiquitin moieties may be masking the LB509 epitope; ***, modified form of α-syn, possibly dimerized α-syn). E: Ubiquitinated α-syn from DLB brain can be deubiquitinated by UCH-L1 in vitro . SDS-soluble fraction from the cingulate cortex of NL-1 or DLB-1 was untreated or reacted with 50 nmol/L of UCH-L1. The samples were analyzed by Western blot analysis using LB509. R represents a lane loaded with 100 ng of recombinant human α-syn. The mobility of molecular mass markers (kd) is depicted on the left of each panel.

    Journal: The American Journal of Pathology

    Article Title: Ubiquitination of ?-Synuclein Is Not Required for Formation of Pathological Inclusions in ?-Synucleinopathies

    doi:

    Figure Lengend Snippet: Insoluble α-syn in diseased brain is ubiquitinated. A: Western blot analysis of biochemically fractionated cingulate cortex from a patient with DLB (DLB-3). Immunoblots were developed with anti-α-syn antibodies LB509 and Syn208 as well as anti-ubiquitin antibody mAb 1510. Twenty μl of LS fraction ( lane 1 ), TX fraction ( lane 2 ), sarkosyl-soluble fraction ( lane 3 ), and SDS-soluble fraction ( lane 4 ) were loaded in separate lanes of 15% SDS-polyacrylamide gels. Note that the SDS-soluble fraction is four times as concentrated as each of the other fractions in that 2.5 ml/g of SDS buffer was used for tissue extraction versus 10 ml/g for each of the other fractions. Arrowhead indicates α-syn monomer (αS) and bracket indicates ubiquitin monomer (Ub). Arrows depict mono-, di-, and tri-ubiquitinated forms of α-syn. B: Western blot analysis (with antibodies Syn208 and mAb 1510) of the SDS-soluble fraction from the cingulate cortex of normal brains (NL-1, NL-2) and DLB brains (DLB-1, DLB-2, and DLB-3). Twenty μl of SDS-soluble fraction was loaded in each lane of a 15% gel. One hundred ng of recombinant human α-syn was loaded in the indicated lanes. C: Western blot analysis of the SDS-soluble fraction from the cingulate cortex of case DLB-1 using various anti-α-syn (LB509, Syn102, Syn211) and anti-ubiquitin (mAb 1510, Conj8) antibodies. Arrows depict mono-, di-, and tri-ubiquitinated forms of α-syn. D: Immunoprecipitation followed by Western blot analysis. α-Syn in the SDS-soluble fraction of the cingulate cortex of DLB-1 was isolated by immunoprecipitation with anti-α-syn antibodies. The sample was analyzed by Western blot analysis using anti-α-syn antibody LB509 and anti-ubiquitin antibody mAb 1510 ( arrows , mono- and di-ubiquitinated forms of α-syn; * and **, possible ubiquitinated forms of α-syn in which ubiquitin moieties may be masking the LB509 epitope; ***, modified form of α-syn, possibly dimerized α-syn). E: Ubiquitinated α-syn from DLB brain can be deubiquitinated by UCH-L1 in vitro . SDS-soluble fraction from the cingulate cortex of NL-1 or DLB-1 was untreated or reacted with 50 nmol/L of UCH-L1. The samples were analyzed by Western blot analysis using LB509. R represents a lane loaded with 100 ng of recombinant human α-syn. The mobility of molecular mass markers (kd) is depicted on the left of each panel.

    Article Snippet: SDS-soluble fractions from the cingulate cortex of neuropathologically normal (NL) or DLB brains were diluted 40-fold in UCH buffer (50 mmol/L HEPES, pH 7.8, 0.5 mmol/L EDTA, 1 mmol/L dithiothreitol) and then concentrated 40-fold using a MicroconYM-10 (Millipore Corp., Bedford, MA) to remove SDS.

    Techniques: Western Blot, Recombinant, Immunoprecipitation, Isolation, Modification, In Vitro

    Erythropoietin Receptor-Mediated Activation of the JAK2/STAT5, RAS/MAPK, and PI3K/AKT Signaling Pathways is Attenuated by BCR-ABL Kinase Activity in K562 Cells A. Normalized hEPO-mediated (10′ stimulation) JAK2 activation in K562 cells treated for 1hr with either 100nM dasatinib or 1μM imatinib. Data represents the average ± SD (n=3). JAK2 activation was monitored by immunoprecipitation and activation loop (Y1007) phosphorylation. JAK2 activation was normalized to the level of phospho-Y1007 observed in the “DMSO” condition for each experimental replicate. B. Upper: Line diagram representation of duration of serum starve, kinase inhibitor treatment, and growth-factor stimulation. Lower: Western immunoblot analysis of whole cell lysates from K562 cells after short-term and prolonged BCR-ABL inhibition (100nM dasatinib: 2hrs, 4hrs, 8hrs, 24hrs; 0.2% DMSO, 24hrs) followed by hEPO stimulation (10′). C. Normalized hEPO-mediated JAK2 activation in K562 cells after short-term (1hr) and prolonged (24hrs) 100nM dasatinib treatment followed hEPO stimulation (10′). Data is representative of triplicate experimental analysis. JAK2 activation and normalization was performed as in (A). D. Normalized RAS-GTP loading in K562 cells after short-term (1hr) and prolonged (24hrs) 100nM dasatinib treatment followed by hEPO stimulation (10′). Data is representative of triplicate experimental analysis. RAS activation was monitored using a RAS-GTP pulldown assay and RAS-GTP levels were normalized to the “DMSO” condition. E. Normalized hEPO-mediated (10′ stimulation) JAK2 activation in K562 cells pretreated for 24hrs with 0.2% DMSO, 100nM dasatinib, 1μM imatinib, 500nM TG101348, dasatinib/TG101348, or imatinib/TG101348. JAK2 activation was monitored and normalized as in (A). F. Upper: Line diagram representation of duration of serum starve, kinase inhibitor treatment, and growth-factor stimulation. Lower: Western immunoblot analysis of whole cell lysates in K562 cells pretreated for 24hrs with 0.2% DMSO, 100nM dasatinib, 1μM imatinib, 500nM TG101348, dasatinib/TG101348, or imatinib/TG101348 followed by hEPO stimulation (10′).

    Journal: Cancer discovery

    Article Title: MEK-Dependent Negative Feedback Underlies BCR-ABL-Mediated Oncogene Addiction

    doi: 10.1158/2159-8290.CD-13-0235

    Figure Lengend Snippet: Erythropoietin Receptor-Mediated Activation of the JAK2/STAT5, RAS/MAPK, and PI3K/AKT Signaling Pathways is Attenuated by BCR-ABL Kinase Activity in K562 Cells A. Normalized hEPO-mediated (10′ stimulation) JAK2 activation in K562 cells treated for 1hr with either 100nM dasatinib or 1μM imatinib. Data represents the average ± SD (n=3). JAK2 activation was monitored by immunoprecipitation and activation loop (Y1007) phosphorylation. JAK2 activation was normalized to the level of phospho-Y1007 observed in the “DMSO” condition for each experimental replicate. B. Upper: Line diagram representation of duration of serum starve, kinase inhibitor treatment, and growth-factor stimulation. Lower: Western immunoblot analysis of whole cell lysates from K562 cells after short-term and prolonged BCR-ABL inhibition (100nM dasatinib: 2hrs, 4hrs, 8hrs, 24hrs; 0.2% DMSO, 24hrs) followed by hEPO stimulation (10′). C. Normalized hEPO-mediated JAK2 activation in K562 cells after short-term (1hr) and prolonged (24hrs) 100nM dasatinib treatment followed hEPO stimulation (10′). Data is representative of triplicate experimental analysis. JAK2 activation and normalization was performed as in (A). D. Normalized RAS-GTP loading in K562 cells after short-term (1hr) and prolonged (24hrs) 100nM dasatinib treatment followed by hEPO stimulation (10′). Data is representative of triplicate experimental analysis. RAS activation was monitored using a RAS-GTP pulldown assay and RAS-GTP levels were normalized to the “DMSO” condition. E. Normalized hEPO-mediated (10′ stimulation) JAK2 activation in K562 cells pretreated for 24hrs with 0.2% DMSO, 100nM dasatinib, 1μM imatinib, 500nM TG101348, dasatinib/TG101348, or imatinib/TG101348. JAK2 activation was monitored and normalized as in (A). F. Upper: Line diagram representation of duration of serum starve, kinase inhibitor treatment, and growth-factor stimulation. Lower: Western immunoblot analysis of whole cell lysates in K562 cells pretreated for 24hrs with 0.2% DMSO, 100nM dasatinib, 1μM imatinib, 500nM TG101348, dasatinib/TG101348, or imatinib/TG101348 followed by hEPO stimulation (10′).

    Article Snippet: Cells were lysed in JAK2 IP buffer (50mM Tris pH 7.6, 100mM NaCl, 1mM EDTA, 1mM EGTA, 0.5% NP40, 0.1% Triton) or RAS IP buffer (50mM Tris pH 7.5, 125mM NaCl, 6.5mM MgCl2 , 5% glycerol, 0.2% NP40) supplemented with 1% protease and 1% phosphatase inhibitors (Calbiochem).

    Techniques: Activation Assay, Activity Assay, Immunoprecipitation, Western Blot, Inhibition

    Global Gene Expression Analysis of Dasatinib Treated K562 Cells Identifies Candidate Mediators Responsible for the Attenuation of Myeloid GF-R Signaling in CML Cells A. ). The heat map at the bottom highlights a few of the genes with increased expression following dasatinib treatment in K562 cells. B. Quantitative PCR (qPCR) analysis of potential negative feedback genes in K562 cells treated with dasatinib (100nM), imatinib (1μM), or PD0325901 (500nM). The average fold expression change (2 (−ΔΔCt) ) and standard deviation (SD fold-change ) for each gene is represented (n=3). C. Upper: Line diagram representation of duration of serum starve, kinase inhibitor treatment, and growth-factor stimulation. Lower: Western immunoblot analysis of RAS and ERK activity before and after hEPO-stimulation (10′) in K562 cells pretreated for 24hrs with 0.2% DMSO, 100nM dasatinib, 500nM PD0325901, or dasatinib/PD0325901. RAS activity was monitored using a RAS-GTP pulldown assay. D. Upper: Line diagram representation of duration of serum starve, kinase inhibitor treatment, and growth-factor stimulation. Lower: Western immunoblot analysis of JAK2 and STAT5 activity before and after hEPO-stimulation in K562 cells treated under the same experimental conditions as (A). JAK2 activation was determined by immunoprecipitation and activation loop (Y1007) phosphorylation.

    Journal: Cancer discovery

    Article Title: MEK-Dependent Negative Feedback Underlies BCR-ABL-Mediated Oncogene Addiction

    doi: 10.1158/2159-8290.CD-13-0235

    Figure Lengend Snippet: Global Gene Expression Analysis of Dasatinib Treated K562 Cells Identifies Candidate Mediators Responsible for the Attenuation of Myeloid GF-R Signaling in CML Cells A. ). The heat map at the bottom highlights a few of the genes with increased expression following dasatinib treatment in K562 cells. B. Quantitative PCR (qPCR) analysis of potential negative feedback genes in K562 cells treated with dasatinib (100nM), imatinib (1μM), or PD0325901 (500nM). The average fold expression change (2 (−ΔΔCt) ) and standard deviation (SD fold-change ) for each gene is represented (n=3). C. Upper: Line diagram representation of duration of serum starve, kinase inhibitor treatment, and growth-factor stimulation. Lower: Western immunoblot analysis of RAS and ERK activity before and after hEPO-stimulation (10′) in K562 cells pretreated for 24hrs with 0.2% DMSO, 100nM dasatinib, 500nM PD0325901, or dasatinib/PD0325901. RAS activity was monitored using a RAS-GTP pulldown assay. D. Upper: Line diagram representation of duration of serum starve, kinase inhibitor treatment, and growth-factor stimulation. Lower: Western immunoblot analysis of JAK2 and STAT5 activity before and after hEPO-stimulation in K562 cells treated under the same experimental conditions as (A). JAK2 activation was determined by immunoprecipitation and activation loop (Y1007) phosphorylation.

    Article Snippet: Cells were lysed in JAK2 IP buffer (50mM Tris pH 7.6, 100mM NaCl, 1mM EDTA, 1mM EGTA, 0.5% NP40, 0.1% Triton) or RAS IP buffer (50mM Tris pH 7.5, 125mM NaCl, 6.5mM MgCl2 , 5% glycerol, 0.2% NP40) supplemented with 1% protease and 1% phosphatase inhibitors (Calbiochem).

    Techniques: Expressing, Real-time Polymerase Chain Reaction, Standard Deviation, Western Blot, Activity Assay, Activation Assay, Immunoprecipitation