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Amresco ethylenediaminetetraacetic acid
Ethylenediaminetetraacetic Acid, supplied by Amresco, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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ethylenediaminetetraacetic acid - by Bioz Stars, 2020-03
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Centrifugation:

Article Title: Molecular characterization of amikacin, kanamycin and capreomycin resistance in M/XDR-TB strains isolated in Thailand
Article Snippet: The cells were resuspended in 400 μl of Tris-EDTA-Tween-lysozyme solution (10 mM Tris-HCl (pH 8.0), 1 mM EDTA, 0.5% (v/v) Tween 80, 2 mg/ml lysozyme (Amresco, USA)), and the mixtures were then incubated at 37°C for 3 h. SDS and proteinase K were added to the cell suspension to generate final concentrations of 1% (w/v) and 1 mg/ml, respectively, prior to incubation at 37°C for 1 h. Then, 80 μl of 5 M NaCl and 80 μl of 10% (w/v) cetyl trimethyl ammonium bromide (CTAB) (Sigma, USA) were added to the suspension, and the suspension was immediately heated at 65°C for 15 min. An equal volume of chloroform-isoamyl alcohol (24:1) (v/v) was added to the suspension. .. The aqueous DNA phase was separated by centrifugation at 12,000xg for 5 min and mixed again with an equal volume of chloroform-isoamyl alcohol (24:1) (v/v).

Article Title: Mammary Fat of Breast Cancer: Gene Expression Profiling and Functional Characterization
Article Snippet: Total tissue protein was extracted with RIPA buffer (50 mM Tris-HCl, pH 7.4, 150 mM NaCl, 1% (v/v) Nonidet-P40, 1 mM EDTA, 1 mM NaF, 10 µg/mL aprotinin, 10 µM leupeptin, and 1 mM phenylmethanesulfonyl fluoride (PMSF), Amresco, PA, USA) and allowed to stand on ice for 30 min to permit lysis. .. Total tissue protein was extracted with RIPA buffer (50 mM Tris-HCl, pH 7.4, 150 mM NaCl, 1% (v/v) Nonidet-P40, 1 mM EDTA, 1 mM NaF, 10 µg/mL aprotinin, 10 µM leupeptin, and 1 mM phenylmethanesulfonyl fluoride (PMSF), Amresco, PA, USA) and allowed to stand on ice for 30 min to permit lysis.

Positive Control:

Article Title: Targeting an Essential GTPase Obg for the Development of Broad-Spectrum Antibiotics
Article Snippet: In addition to the wells with complete reaction mixture (RM) containing 5 μM ObgGC , 250 μM GTP in buffer A (columns 2–22), each plate accommodated a background control comprising of RM lacking ObgGC , (column 1) and a positive control (RM without MgCl2 ; columns 23–24). .. The effect of compounds that may act as Mg2+ chelators and disrupt or inhibit the ObgGC GTPase activity was tested using common chelators including EDTA (Amresco, Solon, OH), ethylene glycol tetra acetic acid (EGTA; Alfa Aesar, Ward Hill, MA), Nitrilotriacetic acid (NTA; TCI, Portland, OR), and citric acid (CA; Sigma Aldrich, St. Louis, MO) at the test-compound concentration used in HTS (40 μM).

MTT Assay:

Article Title: Ferroptosis in Neurons and Cancer Cells Is Similar But Differentially Regulated by Histone Deacetylase Inhibitors
Article Snippet: Laminin (CC095), GSK’872 (530389), Necrostatin-1 inactive (480066), Necrosulfonamide (480073) mouse tumor necrosis factor-α (GF027), U0124 (662006), and rabbit anti-acetylated Histone H4 (1:5000; 06-866; RRID: AB_310270 ) antibody were from Millipore; Boc-DON-Gln-Ile-Val-OMe (B003), 1,3-dimethyl-4,5-diphenyl-2-[(2-oxopropyl)thio]imidazolium trifluorosulfonic acid salt (D004) from Zedira GmbH; 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT assay, G4100) from Promega; erastin (S7242) and entinostat (MS-275, S1053) from Selleck Chemicals. .. EDTA (E177) was obtained from Amresco.

Cytometry:

Article Title: Angiopoietin-2 attenuates angiotensin II-induced aortic aneurysm and atherosclerosis in apolipoprotein E-deficient mice
Article Snippet: Paragraph title: Flow cytometry ... Bone marrow cells were harvested by inserting a 25 gauge needle into the left femur and injecting 500 μl of Dulbecco’s Phosphate Buffered Saline without Ca2+ and Mg2+ (DPBS, Gibco) containing 0.2% BSA and 1 mM EDTA (Amresco).

Enzyme-linked Immunosorbent Assay:

Article Title: Peripheral complement interactions with amyloid β peptide (Aβ) in Alzheimer’s disease: 2. Relationship to Aβ immunotherapy
Article Snippet: NHS plus 10 mM EDTA (Amresco, Solon, OH) (final concentration in the serum) was employed as a control. .. C3a production, one of several standard measures of complement activation, was assayed by ELISA (Affymetrix, Santa Clara, CA, #BHS2089) following the manufacturer’s protocol.

Article Title: Curcumin suppresses epithelial-to-mesenchymal transition of peritoneal mesothelial cells (HMrSV5) through regulation of transforming growth factor-activated kinase 1 (TAK1)
Article Snippet: Trypsin (0.25%) and EDTA (0.02%) were purchased from Amresco (USA). .. A human TGF-β1 ELISA kit was purchased from PeproTech (USA).

Incubation:

Article Title: Sorting and identification of side population cells in the human cervical cancer cell line HeLa
Article Snippet: HeLa cells were maintained as adherent monolayer cultures in high-glucose Dulbecco Modified Eagle's Medium (DMEM, HyClone, USA) supplemented with 10% fetal bovine serum (FBS, HyClone, USA) and incubated at 37°C with 5% CO2 . .. At 80–90% confluency, the cells were washed twice with PBS, and then digested with 0.25% trypsin (Sigma Aldrich, USA) and 0.02% EDTA (Amresco, USA) (v/v, 1:3).

Article Title: Molecular characterization of amikacin, kanamycin and capreomycin resistance in M/XDR-TB strains isolated in Thailand
Article Snippet: .. The cells were resuspended in 400 μl of Tris-EDTA-Tween-lysozyme solution (10 mM Tris-HCl (pH 8.0), 1 mM EDTA, 0.5% (v/v) Tween 80, 2 mg/ml lysozyme (Amresco, USA)), and the mixtures were then incubated at 37°C for 3 h. SDS and proteinase K were added to the cell suspension to generate final concentrations of 1% (w/v) and 1 mg/ml, respectively, prior to incubation at 37°C for 1 h. Then, 80 μl of 5 M NaCl and 80 μl of 10% (w/v) cetyl trimethyl ammonium bromide (CTAB) (Sigma, USA) were added to the suspension, and the suspension was immediately heated at 65°C for 15 min. An equal volume of chloroform-isoamyl alcohol (24:1) (v/v) was added to the suspension. .. The aqueous DNA phase was separated by centrifugation at 12,000xg for 5 min and mixed again with an equal volume of chloroform-isoamyl alcohol (24:1) (v/v).

Article Title: Peripheral complement interactions with amyloid β peptide (Aβ) in Alzheimer’s disease: 2. Relationship to Aβ immunotherapy
Article Snippet: Various concentrations of Aβ prepared in 100 mM Tris, as above, were incubated with either Aβ antibody (4G8, Biolegend, San Diego, CA) or PBS, pH 7.2, after which the solutions were mixed with normal human serum (NHS) (CompTech, Tyler, Texas) for 30 minutes at 37°C. .. NHS plus 10 mM EDTA (Amresco, Solon, OH) (final concentration in the serum) was employed as a control.

Article Title: Targeting an Essential GTPase Obg for the Development of Broad-Spectrum Antibiotics
Article Snippet: ObgGC was incubated with various concentrations of dimethyl sulfoxide (DMSO), 0, 0.5, 1, 2, 3, 4, and 5%, in RM on 384 well plates as outlined above to determine if DMSO interferes with Obg-mediated GTP hydrolysis. .. The effect of compounds that may act as Mg2+ chelators and disrupt or inhibit the ObgGC GTPase activity was tested using common chelators including EDTA (Amresco, Solon, OH), ethylene glycol tetra acetic acid (EGTA; Alfa Aesar, Ward Hill, MA), Nitrilotriacetic acid (NTA; TCI, Portland, OR), and citric acid (CA; Sigma Aldrich, St. Louis, MO) at the test-compound concentration used in HTS (40 μM).

Article Title: The Effect of Heparin-VEGF Multilayer on the Biocompatibility of Decellularized Aortic Valve with Platelet and Endothelial Progenitor Cells
Article Snippet: .. After being incubated for 24 h with D-Hanks containing antibiotics to limit the growth of bacteria (Cefazolin 1.0 mg/ml, Gentamicin 0.4 mg/ml, Amphotericin B 0.5 mg/ml, and Metronidazol 1.2 mg/ml), the valve was incubated with D-Hanks solution with 0.5% Triton X-100 (0694, Amresco), 0.5% sodium deoxycholate, and 0.02% EDTA (0245, Amresco) at 4°C for 48 h under continuous shaking. .. Then, the valve was incubated with ribonuclease A (20 mg/ml) (9001994, Sigma-Aldrich) and deoxyribonuclease (0.2 mg/ml) (DN25, Sigma-Aldrich) at 37°C for 2 h to remove cellular components .

Article Title: Mammary Fat of Breast Cancer: Gene Expression Profiling and Functional Characterization
Article Snippet: Total tissue protein was extracted with RIPA buffer (50 mM Tris-HCl, pH 7.4, 150 mM NaCl, 1% (v/v) Nonidet-P40, 1 mM EDTA, 1 mM NaF, 10 µg/mL aprotinin, 10 µM leupeptin, and 1 mM phenylmethanesulfonyl fluoride (PMSF), Amresco, PA, USA) and allowed to stand on ice for 30 min to permit lysis. .. The membrane was blocked with 1% (w/v) BSA (Amresco, PA, USA) in TBST (10 mM Tris-HCl, pH 7.8, 150 mM NaCl and 0.1% Tween-20) for 2 h and then incubated with a goat anti-human FABP4 primary antibody (1∶200, Santa Cruze, CA, USA), a rabbit anti-human HSL primary antibody (1∶200, Santa Cruze, CA, USA) or a rabbit anti-human UCP1 primary antibody (1∶1000, Abcam, MA, USA) in TBST containing 1% (w/v) BSA overnight at 4°C.

Activity Assay:

Article Title: Targeting an Essential GTPase Obg for the Development of Broad-Spectrum Antibiotics
Article Snippet: .. The effect of compounds that may act as Mg2+ chelators and disrupt or inhibit the ObgGC GTPase activity was tested using common chelators including EDTA (Amresco, Solon, OH), ethylene glycol tetra acetic acid (EGTA; Alfa Aesar, Ward Hill, MA), Nitrilotriacetic acid (NTA; TCI, Portland, OR), and citric acid (CA; Sigma Aldrich, St. Louis, MO) at the test-compound concentration used in HTS (40 μM). ..

Mass Spectrometry:

Article Title: Postoperative recurrence of epithelial ovarian cancer patients and chemoresistance related protein analyses
Article Snippet: Reagents and instruments Urea, thiourea, dithiothreitol (DTT), 3-[3-(cholamidopropyl) dimethylamino] propanesulfonic acid salt (CHAPS), iodoacetamide, tricarboxylaminomethane (Tris), glycine and low melting point agar (10 cm) were from GE Corporation (Shanghai, China); sugar, trichloroacetic acid (TCA), ammonium persulfate (APS), sodium carbonate, N, N, N ‘, N’-tetramethyldiethylamine (TEMED), silver nitrate sodium thiosulfate and EDTA were from Amresco (Washington DC, USA); DC protein assay kit was from Bio-Rad (CA, USA). .. Low temperature high speed centrifuger (TTICH) was from Heraeus (Hanau, Germany); ultraviolet spectrophotometer was from Beckman (Bernard, FL, USA), eppendorf refrigerated centrifuge was from Eppendorf (German); IPGhor isoelectric focusing instrument, DALT-SIX SDS (TTICH type) and ImageScanner Scanner were from Shanghai GE Healthcare; PDquest Analysis Software was from Bio-Rad; ABI 5800 MALDI-TOF/TOF Tandem Mass Spectrometer was from ABI (USA).

Article Title: Ferroptosis in Neurons and Cancer Cells Is Similar But Differentially Regulated by Histone Deacetylase Inhibitors
Article Snippet: Laminin (CC095), GSK’872 (530389), Necrostatin-1 inactive (480066), Necrosulfonamide (480073) mouse tumor necrosis factor-α (GF027), U0124 (662006), and rabbit anti-acetylated Histone H4 (1:5000; 06-866; RRID: AB_310270 ) antibody were from Millipore; Boc-DON-Gln-Ile-Val-OMe (B003), 1,3-dimethyl-4,5-diphenyl-2-[(2-oxopropyl)thio]imidazolium trifluorosulfonic acid salt (D004) from Zedira GmbH; 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT assay, G4100) from Promega; erastin (S7242) and entinostat (MS-275, S1053) from Selleck Chemicals. .. EDTA (E177) was obtained from Amresco.

BIA-KA:

Article Title: Mammary Fat of Breast Cancer: Gene Expression Profiling and Functional Characterization
Article Snippet: Total tissue protein was extracted with RIPA buffer (50 mM Tris-HCl, pH 7.4, 150 mM NaCl, 1% (v/v) Nonidet-P40, 1 mM EDTA, 1 mM NaF, 10 µg/mL aprotinin, 10 µM leupeptin, and 1 mM phenylmethanesulfonyl fluoride (PMSF), Amresco, PA, USA) and allowed to stand on ice for 30 min to permit lysis. .. Total tissue protein was extracted with RIPA buffer (50 mM Tris-HCl, pH 7.4, 150 mM NaCl, 1% (v/v) Nonidet-P40, 1 mM EDTA, 1 mM NaF, 10 µg/mL aprotinin, 10 µM leupeptin, and 1 mM phenylmethanesulfonyl fluoride (PMSF), Amresco, PA, USA) and allowed to stand on ice for 30 min to permit lysis.

Modification:

Article Title: Sorting and identification of side population cells in the human cervical cancer cell line HeLa
Article Snippet: HeLa cells were maintained as adherent monolayer cultures in high-glucose Dulbecco Modified Eagle's Medium (DMEM, HyClone, USA) supplemented with 10% fetal bovine serum (FBS, HyClone, USA) and incubated at 37°C with 5% CO2 . .. At 80–90% confluency, the cells were washed twice with PBS, and then digested with 0.25% trypsin (Sigma Aldrich, USA) and 0.02% EDTA (Amresco, USA) (v/v, 1:3).

Western Blot:

Article Title: Mammary Fat of Breast Cancer: Gene Expression Profiling and Functional Characterization
Article Snippet: Paragraph title: Protein extraction and western blotting ... Total tissue protein was extracted with RIPA buffer (50 mM Tris-HCl, pH 7.4, 150 mM NaCl, 1% (v/v) Nonidet-P40, 1 mM EDTA, 1 mM NaF, 10 µg/mL aprotinin, 10 µM leupeptin, and 1 mM phenylmethanesulfonyl fluoride (PMSF), Amresco, PA, USA) and allowed to stand on ice for 30 min to permit lysis.

Live Dead Assay:

Article Title: Ferroptosis in Neurons and Cancer Cells Is Similar But Differentially Regulated by Histone Deacetylase Inhibitors
Article Snippet: EDTA (E177) was obtained from Amresco. .. DMEM (11965118), DMEM/nutrient mixture F-12 (10565-018), MEM GlutaMAX Supplement (41090101), MEM non-essential amino acids (11140050), fetal bovine serum (16140071), horse serum (26050088), penicillin-streptomycin (15140163), live/dead assay (L3224), NuPAGE 4% and 12% Bis-Tris protein gels (NP0335 and NP0336), MES SDS Running Buffer (NP0002), TaqMan c-Myc (Hs00153408), p21 (Hs00355782), HDAC1 (Hs00606262_g1), HDAC2 (Hs00231032_m1), HDAC3 (Hs00187320_m1), HDAC8 (Hs00954353_g1), GAPDH (4332649) human primers, HDAC1 (Mm02391771), HDAC2 (Mm00515108), HDAC3 (Mm00515916), HDAC8 (Mm01224980_m1) mouse primers, mouse β-actin endogenous control VIC (4352341E), TaqMan RNA-to-CT 1-Step Kit (4392656), and MicroAmp 96-well Reaction Plates (4346906) were purchased from ThermoFisher Scientific.

High Performance Liquid Chromatography:

Article Title: CRISPR/Cas9-based genome-wide screening of Toxoplasma gondii
Article Snippet: .. Blunt needles (SAI Infusion cat. no. B27–50) can be used as an alternative. pU6-SAG1 (Addgene cat. no. 80322) Lourido Toxoplasma CRISPR Library V1 (Addgene cat. no. 80636) pU6-DHFR (Addgene cat. no. 80329) pCas9-CAT (Addgene cat. no. 80323) pU6-Decoy (Addgene cat. no. 80324) Ampicillin (Sigma-Aldrich-Aldrich cat. no. A0166–25G) Glycerol (EMD cat. no. GX0185–6) Methanol-Free Formaldehyde (Thermo Scientific cat. no. 28908) Prolong Gold (Invitrogen cat. no. P1044) Molecular biology-grade water (e.g., from an EMD Millipore Milii-Q Integral System, Fisher Scientific cat. no. ZRXQ003US) Gentamicin (Gibco cat. no. 15710–064) EGTA (Sigma-Aldrich cat. no. E4378–500G) HEPES (Sigma-Aldrich cat. no. 3375–1KG) KH2 PO4 (Sigma-Aldrich cat. no. P5655) K2 HPO4 (Sigma-Aldrich cat. no. 1551128) KCl (Fisher Scientific cat. no. SP138–500) MgCl2 (Mallinckrodt cat. no. 5958) Methanol for HPLC gradient analysis (Millipore, cat. No. 67–56-1) EDTA (Amresco cat. no. 0322–500G) Tris base (Sigma-Aldrich cat. no. TRIS-RO) Glacial acetic acid (Spectrum cat. no. A1010) CAUTION Glacial acetic acid is corrosive; avoid contact with eyes, skin, and clothing; wear protective gloves while handling. .. Na2 HPO4 (Sigma-Aldrich cat. no. 255793) NaCl (Sigma Aldrich cat. no. S7653) Glutathione (Sigma-Aldrich cat. no. G6013–5G) Adenosine Triphosphate (ATP, Sigma-Aldrich cat. no. A6419–1G) CaCl2 (Sigma-Aldrich cat. no. C5080–1KG) Chloramphenicol (Sigma-Aldrich cat. no. C3175–100MG) Pyrimethamine (Sigma-Aldrich cat. no. 46706–250MG) NaOH (Sigma-Aldrich cat. no. S2770) CAUTION Sodium hydroxide is a strong base that can cause severe skin burns and eye injuries; avoid contact with eyes, skin and clothing; use in a fume hood and wear protective eyewear and gloves during handling.

Flow Cytometry:

Article Title: Arrestin Expression in E. coli and Purification
Article Snippet: .. Luria Broth (LB) (Sigma; prepare according to the manufacturer’s instructions) 100 mg/ml ampicillin stock solution in distilled water (store at 4°C) 100 mM Isopropyl-β-D-thiogalactopyranoside stock solution (IPTG) in distilled water (store at −80°C) Glycerol (Sigma) 1.0 M Tris-HCl buffer solution, pH 8.0 (Amresco) 0.5 M Ethylene glycol tetraacetic acid solution (EGTA, BioWorld) 1.0 M Glucose stock solution in distilled water (store at −20°C) Phenylmethylsulfonyl fluoride (PMSF) Dimethyl sulfoxide (DMSO) 1.0 M Phenylmethanesulfonyl fluoride (PMSF) (prepare a fresh solution in water-free DMSO) 1.0 M Benzamidine (BA) stock solution in distilled water (store at −80°C) 1.0 M Magnesium chloride solution (Sigma) 5.0 M Sodium Chloride solution (Sigma) 4.0 M Dithiothreitol (DTT) stock solution in distilled water (store at −20°C) 0.5 M Ethylenediaminetetraacetic acid (EDTA, Amresco) 2X SDS Sample buffer (Sigma) Lysozyme from chicken egg white (Sigma) Protease inhibitor cocktail for E. coli (P-8465, Sigma, prepared according to manufacturer’s instructions) 15,000 U Deoxyribonuclease I from bovine pancreas (DNAse I, Sigma) (stock solution: dissolve in 1 ml of Lysis buffer) 20,000 U Deoxyribonuclease II from porcine spleen (DNAse II, Sigma) (stock solution: dissolve in 1 ml of Lysis buffer) 100 mg/ml Ribonuclease A from bovine pancreas (RNAse A, Sigma) (stock solution: dissolve in 1 ml of Lysis buffer) Heparin Sepharose 6 Fast Flow (GE Healthcare) Q Sepharose Fast Flow (GE Healthcare) SP Sepharose Fast Flow (GE Healthcare) .. RC-5B Plus centrifuge (Sorvall) SLA 3000 rotor (Sorvall) TL-100 tabletop ultracentrifuge (Beckman) TLA120.1 rotor (Beckman); Econo-Column Chromatography Columns, 1.5 × 20 cm (Bio-Rad, cat. #737-1522) GradiFrac Chromatography system (Pharmacia Biotech, Sweden) Additional reagents and equipment for performing SDS-PAGE electrophoresis and Western Blotting - described in detail in ( ; )).

Article Title: Angiopoietin-2 attenuates angiotensin II-induced aortic aneurysm and atherosclerosis in apolipoprotein E-deficient mice
Article Snippet: Paragraph title: Flow cytometry ... Bone marrow cells were harvested by inserting a 25 gauge needle into the left femur and injecting 500 μl of Dulbecco’s Phosphate Buffered Saline without Ca2+ and Mg2+ (DPBS, Gibco) containing 0.2% BSA and 1 mM EDTA (Amresco).

Concentration Assay:

Article Title: Molecular characterization of amikacin, kanamycin and capreomycin resistance in M/XDR-TB strains isolated in Thailand
Article Snippet: The cells were resuspended in 400 μl of Tris-EDTA-Tween-lysozyme solution (10 mM Tris-HCl (pH 8.0), 1 mM EDTA, 0.5% (v/v) Tween 80, 2 mg/ml lysozyme (Amresco, USA)), and the mixtures were then incubated at 37°C for 3 h. SDS and proteinase K were added to the cell suspension to generate final concentrations of 1% (w/v) and 1 mg/ml, respectively, prior to incubation at 37°C for 1 h. Then, 80 μl of 5 M NaCl and 80 μl of 10% (w/v) cetyl trimethyl ammonium bromide (CTAB) (Sigma, USA) were added to the suspension, and the suspension was immediately heated at 65°C for 15 min. An equal volume of chloroform-isoamyl alcohol (24:1) (v/v) was added to the suspension. .. RNaseA (Qiagen, Germany) was added to the total nucleic acid solution to generate a final concentration of 0.5 μg/μl, and the tube was subsequently incubated at 37°C for 1 h.

Article Title: Peripheral complement interactions with amyloid β peptide (Aβ) in Alzheimer’s disease: 2. Relationship to Aβ immunotherapy
Article Snippet: .. NHS plus 10 mM EDTA (Amresco, Solon, OH) (final concentration in the serum) was employed as a control. .. C3a production, one of several standard measures of complement activation, was assayed by ELISA (Affymetrix, Santa Clara, CA, #BHS2089) following the manufacturer’s protocol.

Article Title: Targeting an Essential GTPase Obg for the Development of Broad-Spectrum Antibiotics
Article Snippet: .. The effect of compounds that may act as Mg2+ chelators and disrupt or inhibit the ObgGC GTPase activity was tested using common chelators including EDTA (Amresco, Solon, OH), ethylene glycol tetra acetic acid (EGTA; Alfa Aesar, Ward Hill, MA), Nitrilotriacetic acid (NTA; TCI, Portland, OR), and citric acid (CA; Sigma Aldrich, St. Louis, MO) at the test-compound concentration used in HTS (40 μM). ..

Protease Inhibitor:

Article Title: Arrestin Expression in E. coli and Purification
Article Snippet: .. Luria Broth (LB) (Sigma; prepare according to the manufacturer’s instructions) 100 mg/ml ampicillin stock solution in distilled water (store at 4°C) 100 mM Isopropyl-β-D-thiogalactopyranoside stock solution (IPTG) in distilled water (store at −80°C) Glycerol (Sigma) 1.0 M Tris-HCl buffer solution, pH 8.0 (Amresco) 0.5 M Ethylene glycol tetraacetic acid solution (EGTA, BioWorld) 1.0 M Glucose stock solution in distilled water (store at −20°C) Phenylmethylsulfonyl fluoride (PMSF) Dimethyl sulfoxide (DMSO) 1.0 M Phenylmethanesulfonyl fluoride (PMSF) (prepare a fresh solution in water-free DMSO) 1.0 M Benzamidine (BA) stock solution in distilled water (store at −80°C) 1.0 M Magnesium chloride solution (Sigma) 5.0 M Sodium Chloride solution (Sigma) 4.0 M Dithiothreitol (DTT) stock solution in distilled water (store at −20°C) 0.5 M Ethylenediaminetetraacetic acid (EDTA, Amresco) 2X SDS Sample buffer (Sigma) Lysozyme from chicken egg white (Sigma) Protease inhibitor cocktail for E. coli (P-8465, Sigma, prepared according to manufacturer’s instructions) 15,000 U Deoxyribonuclease I from bovine pancreas (DNAse I, Sigma) (stock solution: dissolve in 1 ml of Lysis buffer) 20,000 U Deoxyribonuclease II from porcine spleen (DNAse II, Sigma) (stock solution: dissolve in 1 ml of Lysis buffer) 100 mg/ml Ribonuclease A from bovine pancreas (RNAse A, Sigma) (stock solution: dissolve in 1 ml of Lysis buffer) Heparin Sepharose 6 Fast Flow (GE Healthcare) Q Sepharose Fast Flow (GE Healthcare) SP Sepharose Fast Flow (GE Healthcare) .. RC-5B Plus centrifuge (Sorvall) SLA 3000 rotor (Sorvall) TL-100 tabletop ultracentrifuge (Beckman) TLA120.1 rotor (Beckman); Econo-Column Chromatography Columns, 1.5 × 20 cm (Bio-Rad, cat. #737-1522) GradiFrac Chromatography system (Pharmacia Biotech, Sweden) Additional reagents and equipment for performing SDS-PAGE electrophoresis and Western Blotting - described in detail in ( ; )).

Article Title: Ferroptosis in Neurons and Cancer Cells Is Similar But Differentially Regulated by Histone Deacetylase Inhibitors
Article Snippet: Actinomycin D (A1410), Chloroquine (C6628), cycloheximide (01810), Cystamine dihydrochloride (C8707), Deferoxamine (D9533), Ferrostatin-1 (SML0583), Homocysteate (H9633), Mithramycin A (M6891), N -acetylcysteine (A7250), rapamycin (R8781), sodium butyrate (303410), protease inhibitor cocktail (P8340), poly-d -lysine (P6407), poly-l -lysine (P4704), N1 supplement (N6530), collagenase (C9697), EGTA (E0396), sodium orthovanadate (S6508), mouse anti-β-actin (1:20,000; clone AC-74, A5316; RRID: AB_476743 ), and Tween 20 (P7949) were obtained from Sigma-Aldrich. .. EDTA (E177) was obtained from Amresco.

Transferring:

Article Title: CRISPR/Cas9-based genome-wide screening of Toxoplasma gondii
Article Snippet: Eppendorf tubes (Axygen cat. no. MCT-150-C) 50 ml tubes (Corning cat. no. 430828) 15 ml tubes (Corning cat. no. 430052) Pipettes (Fisher Scientific cat. no. F144802G and F167300G) Pipette tips with filters (USA Scientific cat. no. 1120–1810, 1120–8810, 1122–1830 and 1120–3810) Sterile Acrodisc® Syringe Filters with HT Tuffryn® Membrane 0.2 μm, 25 mm (Pall Life Sciences cat. no. 4192). .. Blunt needles (SAI Infusion cat. no. B27–50) can be used as an alternative. pU6-SAG1 (Addgene cat. no. 80322) Lourido Toxoplasma CRISPR Library V1 (Addgene cat. no. 80636) pU6-DHFR (Addgene cat. no. 80329) pCas9-CAT (Addgene cat. no. 80323) pU6-Decoy (Addgene cat. no. 80324) Ampicillin (Sigma-Aldrich-Aldrich cat. no. A0166–25G) Glycerol (EMD cat. no. GX0185–6) Methanol-Free Formaldehyde (Thermo Scientific cat. no. 28908) Prolong Gold (Invitrogen cat. no. P1044) Molecular biology-grade water (e.g., from an EMD Millipore Milii-Q Integral System, Fisher Scientific cat. no. ZRXQ003US) Gentamicin (Gibco cat. no. 15710–064) EGTA (Sigma-Aldrich cat. no. E4378–500G) HEPES (Sigma-Aldrich cat. no. 3375–1KG) KH2 PO4 (Sigma-Aldrich cat. no. P5655) K2 HPO4 (Sigma-Aldrich cat. no. 1551128) KCl (Fisher Scientific cat. no. SP138–500) MgCl2 (Mallinckrodt cat. no. 5958) Methanol for HPLC gradient analysis (Millipore, cat. No. 67–56-1) EDTA (Amresco cat. no. 0322–500G) Tris base (Sigma-Aldrich cat. no. TRIS-RO) Glacial acetic acid (Spectrum cat. no. A1010) CAUTION Glacial acetic acid is corrosive; avoid contact with eyes, skin, and clothing; wear protective gloves while handling.

Cell Culture:

Article Title: Sorting and identification of side population cells in the human cervical cancer cell line HeLa
Article Snippet: Paragraph title: Cell culture ... At 80–90% confluency, the cells were washed twice with PBS, and then digested with 0.25% trypsin (Sigma Aldrich, USA) and 0.02% EDTA (Amresco, USA) (v/v, 1:3).

other:

Article Title: Characterization of intestinal dendritic cells in murine norovirus infection
Article Snippet: The following reagents were used: Isoflurane (NDC 57319-507-06, Phoenix Pharmaceutical, Inc. St. Joseph MO), collagenase I (Worthington Biochemical Corporation, Lakewood NK), hyaluronidase type II (Sigma-Aldrich, St. Louis MO), DNase I (Worthington Biochemical), EDTA (AMRESCO Inc, Solon OH), 4′,6-diamidino-2-phenylindole (DAPI, Thermo Scientific, Rockford IL), ovalbumin (OVA, Sigma-Aldrich), Tissue-Tek OCT compound (Sakura Finetek USA, Inc. Torrance CA), paraformaldehyde (Sigma-Aldrich); Fluoromount G (EMS, Hatfield PA); Trizol (Invitrogen Corporation, Carlsbad CA).

Article Title: MiR199b Suppresses Expression of Hypoxia-Inducible Factor 1? (HIF-1?) in Prostate Cancer Cells
Article Snippet: Tris base, Tween 20, DTT, and EDTA were from Amresco.

Protein Concentration:

Article Title: Mammary Fat of Breast Cancer: Gene Expression Profiling and Functional Characterization
Article Snippet: Total tissue protein was extracted with RIPA buffer (50 mM Tris-HCl, pH 7.4, 150 mM NaCl, 1% (v/v) Nonidet-P40, 1 mM EDTA, 1 mM NaF, 10 µg/mL aprotinin, 10 µM leupeptin, and 1 mM phenylmethanesulfonyl fluoride (PMSF), Amresco, PA, USA) and allowed to stand on ice for 30 min to permit lysis. .. Total tissue protein was extracted with RIPA buffer (50 mM Tris-HCl, pH 7.4, 150 mM NaCl, 1% (v/v) Nonidet-P40, 1 mM EDTA, 1 mM NaF, 10 µg/mL aprotinin, 10 µM leupeptin, and 1 mM phenylmethanesulfonyl fluoride (PMSF), Amresco, PA, USA) and allowed to stand on ice for 30 min to permit lysis.

Polymerase Chain Reaction:

Article Title: CRISPR/Cas9-based genome-wide screening of Toxoplasma gondii
Article Snippet: Petri dishes (Falcon cat. no. 351029) Culture tubes (Corning cat. no. 352051) 24-well tissue culture dishes (Costar cat. no. 3524) Parafilm (VWR cat. no. 291–1214) T12.5 tissue culture flasks (Falcon cat. no. 353018) 15 cm tissue culture dishes (Falcon cat. no. 353025) Cell scrapers (Costar cat. no. 3010) PCR tubes (BioRad cat. no. TF102010) Cryotubes (Thermo Scientific cat. no. 5000–0012) 27½ -gauge needles (BD cat. no. 305109) CAUTION Exercise extreme care when handling live T. gondii with sharps. .. Blunt needles (SAI Infusion cat. no. B27–50) can be used as an alternative. pU6-SAG1 (Addgene cat. no. 80322) Lourido Toxoplasma CRISPR Library V1 (Addgene cat. no. 80636) pU6-DHFR (Addgene cat. no. 80329) pCas9-CAT (Addgene cat. no. 80323) pU6-Decoy (Addgene cat. no. 80324) Ampicillin (Sigma-Aldrich-Aldrich cat. no. A0166–25G) Glycerol (EMD cat. no. GX0185–6) Methanol-Free Formaldehyde (Thermo Scientific cat. no. 28908) Prolong Gold (Invitrogen cat. no. P1044) Molecular biology-grade water (e.g., from an EMD Millipore Milii-Q Integral System, Fisher Scientific cat. no. ZRXQ003US) Gentamicin (Gibco cat. no. 15710–064) EGTA (Sigma-Aldrich cat. no. E4378–500G) HEPES (Sigma-Aldrich cat. no. 3375–1KG) KH2 PO4 (Sigma-Aldrich cat. no. P5655) K2 HPO4 (Sigma-Aldrich cat. no. 1551128) KCl (Fisher Scientific cat. no. SP138–500) MgCl2 (Mallinckrodt cat. no. 5958) Methanol for HPLC gradient analysis (Millipore, cat. No. 67–56-1) EDTA (Amresco cat. no. 0322–500G) Tris base (Sigma-Aldrich cat. no. TRIS-RO) Glacial acetic acid (Spectrum cat. no. A1010) CAUTION Glacial acetic acid is corrosive; avoid contact with eyes, skin, and clothing; wear protective gloves while handling.

DC Protein Assay:

Article Title: Postoperative recurrence of epithelial ovarian cancer patients and chemoresistance related protein analyses
Article Snippet: .. Reagents and instruments Urea, thiourea, dithiothreitol (DTT), 3-[3-(cholamidopropyl) dimethylamino] propanesulfonic acid salt (CHAPS), iodoacetamide, tricarboxylaminomethane (Tris), glycine and low melting point agar (10 cm) were from GE Corporation (Shanghai, China); sugar, trichloroacetic acid (TCA), ammonium persulfate (APS), sodium carbonate, N, N, N ‘, N’-tetramethyldiethylamine (TEMED), silver nitrate sodium thiosulfate and EDTA were from Amresco (Washington DC, USA); DC protein assay kit was from Bio-Rad (CA, USA). .. Bovine serum albumin (BSA) was purchased from Pierce; albumin and IgG protein removal and desalting kits were Merck (NJ, USA); trypsin was from Promega (Madison, WI, USA); formic acid, acetonitrile, tris-HCl (pH 6.8, pH 8.5, pH 8.8), coomassie Brilliant blue R-250, coomassie brilliant blue G-250, bromophenol blue, acetone, methanol, phosphoric acid, n-butanol, ethanol, glacial acetic acid, ammonium sulfate were obtained from Sigma (St. Louis, MI, USA).

Article Title: Ferroptosis in Neurons and Cancer Cells Is Similar But Differentially Regulated by Histone Deacetylase Inhibitors
Article Snippet: EDTA (E177) was obtained from Amresco. .. Triton X-100 (161-0407), DC Protein Assay Kit I (5000111), Quick Start Bradford Reagent (500-0205), and Protein Dual Color Standard (161-0374) were purchased from Bio-Rad.

Nucleic Acid Electrophoresis:

Article Title: Mammary Fat of Breast Cancer: Gene Expression Profiling and Functional Characterization
Article Snippet: Total tissue protein was extracted with RIPA buffer (50 mM Tris-HCl, pH 7.4, 150 mM NaCl, 1% (v/v) Nonidet-P40, 1 mM EDTA, 1 mM NaF, 10 µg/mL aprotinin, 10 µM leupeptin, and 1 mM phenylmethanesulfonyl fluoride (PMSF), Amresco, PA, USA) and allowed to stand on ice for 30 min to permit lysis. .. Aliquots (60 µg) of protein were separated on a 12% SDS-polyacrylamide gel electrophoresis and transferred onto a poly (vinylidene difluoride) membrane (Millipore, Boston, USA).

Fluorescence:

Article Title: Effect of Total Flavonoids of Rhizoma drynariae on Tibial Dyschondroplasia by Regulating BMP-2 and Runx2 Expression in Chickens
Article Snippet: EDTA was purchased from Amresco analytical grade. .. Reverse transcription, fluorescence quantitative PCR kits were purchased from Beijing TransGen Biotech.

Isolation:

Article Title: Molecular characterization of amikacin, kanamycin and capreomycin resistance in M/XDR-TB strains isolated in Thailand
Article Snippet: Paragraph title: Isolation of genomic DNA ... The cells were resuspended in 400 μl of Tris-EDTA-Tween-lysozyme solution (10 mM Tris-HCl (pH 8.0), 1 mM EDTA, 0.5% (v/v) Tween 80, 2 mg/ml lysozyme (Amresco, USA)), and the mixtures were then incubated at 37°C for 3 h. SDS and proteinase K were added to the cell suspension to generate final concentrations of 1% (w/v) and 1 mg/ml, respectively, prior to incubation at 37°C for 1 h. Then, 80 μl of 5 M NaCl and 80 μl of 10% (w/v) cetyl trimethyl ammonium bromide (CTAB) (Sigma, USA) were added to the suspension, and the suspension was immediately heated at 65°C for 15 min. An equal volume of chloroform-isoamyl alcohol (24:1) (v/v) was added to the suspension.

Immunohistochemistry:

Article Title: Effect of Total Flavonoids of Rhizoma drynariae on Tibial Dyschondroplasia by Regulating BMP-2 and Runx2 Expression in Chickens
Article Snippet: EDTA was purchased from Amresco analytical grade. .. Ready-to-use SABC immunohistochemical staining kit purchased from Wuhan Boster Company.

Protein Extraction:

Article Title: Mammary Fat of Breast Cancer: Gene Expression Profiling and Functional Characterization
Article Snippet: Paragraph title: Protein extraction and western blotting ... Total tissue protein was extracted with RIPA buffer (50 mM Tris-HCl, pH 7.4, 150 mM NaCl, 1% (v/v) Nonidet-P40, 1 mM EDTA, 1 mM NaF, 10 µg/mL aprotinin, 10 µM leupeptin, and 1 mM phenylmethanesulfonyl fluoride (PMSF), Amresco, PA, USA) and allowed to stand on ice for 30 min to permit lysis.

CRISPR:

Article Title: CRISPR/Cas9-based genome-wide screening of Toxoplasma gondii
Article Snippet: .. Blunt needles (SAI Infusion cat. no. B27–50) can be used as an alternative. pU6-SAG1 (Addgene cat. no. 80322) Lourido Toxoplasma CRISPR Library V1 (Addgene cat. no. 80636) pU6-DHFR (Addgene cat. no. 80329) pCas9-CAT (Addgene cat. no. 80323) pU6-Decoy (Addgene cat. no. 80324) Ampicillin (Sigma-Aldrich-Aldrich cat. no. A0166–25G) Glycerol (EMD cat. no. GX0185–6) Methanol-Free Formaldehyde (Thermo Scientific cat. no. 28908) Prolong Gold (Invitrogen cat. no. P1044) Molecular biology-grade water (e.g., from an EMD Millipore Milii-Q Integral System, Fisher Scientific cat. no. ZRXQ003US) Gentamicin (Gibco cat. no. 15710–064) EGTA (Sigma-Aldrich cat. no. E4378–500G) HEPES (Sigma-Aldrich cat. no. 3375–1KG) KH2 PO4 (Sigma-Aldrich cat. no. P5655) K2 HPO4 (Sigma-Aldrich cat. no. 1551128) KCl (Fisher Scientific cat. no. SP138–500) MgCl2 (Mallinckrodt cat. no. 5958) Methanol for HPLC gradient analysis (Millipore, cat. No. 67–56-1) EDTA (Amresco cat. no. 0322–500G) Tris base (Sigma-Aldrich cat. no. TRIS-RO) Glacial acetic acid (Spectrum cat. no. A1010) CAUTION Glacial acetic acid is corrosive; avoid contact with eyes, skin, and clothing; wear protective gloves while handling. .. Na2 HPO4 (Sigma-Aldrich cat. no. 255793) NaCl (Sigma Aldrich cat. no. S7653) Glutathione (Sigma-Aldrich cat. no. G6013–5G) Adenosine Triphosphate (ATP, Sigma-Aldrich cat. no. A6419–1G) CaCl2 (Sigma-Aldrich cat. no. C5080–1KG) Chloramphenicol (Sigma-Aldrich cat. no. C3175–100MG) Pyrimethamine (Sigma-Aldrich cat. no. 46706–250MG) NaOH (Sigma-Aldrich cat. no. S2770) CAUTION Sodium hydroxide is a strong base that can cause severe skin burns and eye injuries; avoid contact with eyes, skin and clothing; use in a fume hood and wear protective eyewear and gloves during handling.

Activated Clotting Time Assay:

Article Title: Targeting an Essential GTPase Obg for the Development of Broad-Spectrum Antibiotics
Article Snippet: .. The effect of compounds that may act as Mg2+ chelators and disrupt or inhibit the ObgGC GTPase activity was tested using common chelators including EDTA (Amresco, Solon, OH), ethylene glycol tetra acetic acid (EGTA; Alfa Aesar, Ward Hill, MA), Nitrilotriacetic acid (NTA; TCI, Portland, OR), and citric acid (CA; Sigma Aldrich, St. Louis, MO) at the test-compound concentration used in HTS (40 μM). ..

Software:

Article Title: Postoperative recurrence of epithelial ovarian cancer patients and chemoresistance related protein analyses
Article Snippet: Reagents and instruments Urea, thiourea, dithiothreitol (DTT), 3-[3-(cholamidopropyl) dimethylamino] propanesulfonic acid salt (CHAPS), iodoacetamide, tricarboxylaminomethane (Tris), glycine and low melting point agar (10 cm) were from GE Corporation (Shanghai, China); sugar, trichloroacetic acid (TCA), ammonium persulfate (APS), sodium carbonate, N, N, N ‘, N’-tetramethyldiethylamine (TEMED), silver nitrate sodium thiosulfate and EDTA were from Amresco (Washington DC, USA); DC protein assay kit was from Bio-Rad (CA, USA). .. Low temperature high speed centrifuger (TTICH) was from Heraeus (Hanau, Germany); ultraviolet spectrophotometer was from Beckman (Bernard, FL, USA), eppendorf refrigerated centrifuge was from Eppendorf (German); IPGhor isoelectric focusing instrument, DALT-SIX SDS (TTICH type) and ImageScanner Scanner were from Shanghai GE Healthcare; PDquest Analysis Software was from Bio-Rad; ABI 5800 MALDI-TOF/TOF Tandem Mass Spectrometer was from ABI (USA).

Real-time Polymerase Chain Reaction:

Article Title: Effect of Total Flavonoids of Rhizoma drynariae on Tibial Dyschondroplasia by Regulating BMP-2 and Runx2 Expression in Chickens
Article Snippet: EDTA was purchased from Amresco analytical grade. .. Reverse transcription, fluorescence quantitative PCR kits were purchased from Beijing TransGen Biotech.

Spectrophotometry:

Article Title: Postoperative recurrence of epithelial ovarian cancer patients and chemoresistance related protein analyses
Article Snippet: Reagents and instruments Urea, thiourea, dithiothreitol (DTT), 3-[3-(cholamidopropyl) dimethylamino] propanesulfonic acid salt (CHAPS), iodoacetamide, tricarboxylaminomethane (Tris), glycine and low melting point agar (10 cm) were from GE Corporation (Shanghai, China); sugar, trichloroacetic acid (TCA), ammonium persulfate (APS), sodium carbonate, N, N, N ‘, N’-tetramethyldiethylamine (TEMED), silver nitrate sodium thiosulfate and EDTA were from Amresco (Washington DC, USA); DC protein assay kit was from Bio-Rad (CA, USA). .. Low temperature high speed centrifuger (TTICH) was from Heraeus (Hanau, Germany); ultraviolet spectrophotometer was from Beckman (Bernard, FL, USA), eppendorf refrigerated centrifuge was from Eppendorf (German); IPGhor isoelectric focusing instrument, DALT-SIX SDS (TTICH type) and ImageScanner Scanner were from Shanghai GE Healthcare; PDquest Analysis Software was from Bio-Rad; ABI 5800 MALDI-TOF/TOF Tandem Mass Spectrometer was from ABI (USA).

Activation Assay:

Article Title: Peripheral complement interactions with amyloid β peptide (Aβ) in Alzheimer’s disease: 2. Relationship to Aβ immunotherapy
Article Snippet: Paragraph title: 2.03 Serum complement activation ... NHS plus 10 mM EDTA (Amresco, Solon, OH) (final concentration in the serum) was employed as a control.

Lysis:

Article Title: Arrestin Expression in E. coli and Purification
Article Snippet: .. Luria Broth (LB) (Sigma; prepare according to the manufacturer’s instructions) 100 mg/ml ampicillin stock solution in distilled water (store at 4°C) 100 mM Isopropyl-β-D-thiogalactopyranoside stock solution (IPTG) in distilled water (store at −80°C) Glycerol (Sigma) 1.0 M Tris-HCl buffer solution, pH 8.0 (Amresco) 0.5 M Ethylene glycol tetraacetic acid solution (EGTA, BioWorld) 1.0 M Glucose stock solution in distilled water (store at −20°C) Phenylmethylsulfonyl fluoride (PMSF) Dimethyl sulfoxide (DMSO) 1.0 M Phenylmethanesulfonyl fluoride (PMSF) (prepare a fresh solution in water-free DMSO) 1.0 M Benzamidine (BA) stock solution in distilled water (store at −80°C) 1.0 M Magnesium chloride solution (Sigma) 5.0 M Sodium Chloride solution (Sigma) 4.0 M Dithiothreitol (DTT) stock solution in distilled water (store at −20°C) 0.5 M Ethylenediaminetetraacetic acid (EDTA, Amresco) 2X SDS Sample buffer (Sigma) Lysozyme from chicken egg white (Sigma) Protease inhibitor cocktail for E. coli (P-8465, Sigma, prepared according to manufacturer’s instructions) 15,000 U Deoxyribonuclease I from bovine pancreas (DNAse I, Sigma) (stock solution: dissolve in 1 ml of Lysis buffer) 20,000 U Deoxyribonuclease II from porcine spleen (DNAse II, Sigma) (stock solution: dissolve in 1 ml of Lysis buffer) 100 mg/ml Ribonuclease A from bovine pancreas (RNAse A, Sigma) (stock solution: dissolve in 1 ml of Lysis buffer) Heparin Sepharose 6 Fast Flow (GE Healthcare) Q Sepharose Fast Flow (GE Healthcare) SP Sepharose Fast Flow (GE Healthcare) .. RC-5B Plus centrifuge (Sorvall) SLA 3000 rotor (Sorvall) TL-100 tabletop ultracentrifuge (Beckman) TLA120.1 rotor (Beckman); Econo-Column Chromatography Columns, 1.5 × 20 cm (Bio-Rad, cat. #737-1522) GradiFrac Chromatography system (Pharmacia Biotech, Sweden) Additional reagents and equipment for performing SDS-PAGE electrophoresis and Western Blotting - described in detail in ( ; )).

Article Title: Mammary Fat of Breast Cancer: Gene Expression Profiling and Functional Characterization
Article Snippet: .. Total tissue protein was extracted with RIPA buffer (50 mM Tris-HCl, pH 7.4, 150 mM NaCl, 1% (v/v) Nonidet-P40, 1 mM EDTA, 1 mM NaF, 10 µg/mL aprotinin, 10 µM leupeptin, and 1 mM phenylmethanesulfonyl fluoride (PMSF), Amresco, PA, USA) and allowed to stand on ice for 30 min to permit lysis. .. After centrifugation at 11,000 rpm for 10 min at 4°C, protein content in the supernatant was collected and the protein concentration was determined using a BCA assay (Pierce, Rockford, USA).

Staining:

Article Title: Effect of Total Flavonoids of Rhizoma drynariae on Tibial Dyschondroplasia by Regulating BMP-2 and Runx2 Expression in Chickens
Article Snippet: EDTA was purchased from Amresco analytical grade. .. Ready-to-use SABC immunohistochemical staining kit purchased from Wuhan Boster Company.

Hood:

Article Title: CRISPR/Cas9-based genome-wide screening of Toxoplasma gondii
Article Snippet: Blunt needles (SAI Infusion cat. no. B27–50) can be used as an alternative. pU6-SAG1 (Addgene cat. no. 80322) Lourido Toxoplasma CRISPR Library V1 (Addgene cat. no. 80636) pU6-DHFR (Addgene cat. no. 80329) pCas9-CAT (Addgene cat. no. 80323) pU6-Decoy (Addgene cat. no. 80324) Ampicillin (Sigma-Aldrich-Aldrich cat. no. A0166–25G) Glycerol (EMD cat. no. GX0185–6) Methanol-Free Formaldehyde (Thermo Scientific cat. no. 28908) Prolong Gold (Invitrogen cat. no. P1044) Molecular biology-grade water (e.g., from an EMD Millipore Milii-Q Integral System, Fisher Scientific cat. no. ZRXQ003US) Gentamicin (Gibco cat. no. 15710–064) EGTA (Sigma-Aldrich cat. no. E4378–500G) HEPES (Sigma-Aldrich cat. no. 3375–1KG) KH2 PO4 (Sigma-Aldrich cat. no. P5655) K2 HPO4 (Sigma-Aldrich cat. no. 1551128) KCl (Fisher Scientific cat. no. SP138–500) MgCl2 (Mallinckrodt cat. no. 5958) Methanol for HPLC gradient analysis (Millipore, cat. No. 67–56-1) EDTA (Amresco cat. no. 0322–500G) Tris base (Sigma-Aldrich cat. no. TRIS-RO) Glacial acetic acid (Spectrum cat. no. A1010) CAUTION Glacial acetic acid is corrosive; avoid contact with eyes, skin, and clothing; wear protective gloves while handling. .. Na2 HPO4 (Sigma-Aldrich cat. no. 255793) NaCl (Sigma Aldrich cat. no. S7653) Glutathione (Sigma-Aldrich cat. no. G6013–5G) Adenosine Triphosphate (ATP, Sigma-Aldrich cat. no. A6419–1G) CaCl2 (Sigma-Aldrich cat. no. C5080–1KG) Chloramphenicol (Sigma-Aldrich cat. no. C3175–100MG) Pyrimethamine (Sigma-Aldrich cat. no. 46706–250MG) NaOH (Sigma-Aldrich cat. no. S2770) CAUTION Sodium hydroxide is a strong base that can cause severe skin burns and eye injuries; avoid contact with eyes, skin and clothing; use in a fume hood and wear protective eyewear and gloves during handling.

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    Amresco immunoprecipitation lysis buffer
    SIRT6 promotes the ubiquitination of p27 ( A ) 293 cells were transfected with SIRT6 and vector, twenty four hours after transfection, cells were treated with MG132 or acetyl-leu-leu-norleucinal (ALLN) 5 h before harvesting. Proteins were analyzed by western blot. ( B ) An in vivo ubiquitination assay was performed. 293 cells were transiently transfected with SIRT6 and vector, siRNA-SIRT6 and NC (negative control). Forty two hours later, cells were treated with MG132 for 5 h. Cells were lysed and proceeded for <t>co-immunoprecipitation</t> using the p27 antibody.
    Immunoprecipitation Lysis Buffer, supplied by Amresco, used in various techniques. Bioz Stars score: 89/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/immunoprecipitation lysis buffer/product/Amresco
    Average 89 stars, based on 2 article reviews
    Price from $9.99 to $1999.99
    immunoprecipitation lysis buffer - by Bioz Stars, 2020-03
    89/100 stars
      Buy from Supplier

    99
    Amresco edta
    Complement opsonization of Aβ and Aβ/4G8 ICs A) After exposure to <t>NHS</t> to permit complement activation, Aβ alone and Aβ IC solutions were run on SDS reducing/denaturing Western blots to detect the complement opsonin iC3b. <t>EDTA</t> treatment (lanes 2 and 4 of the iC3b blot) abolished complement activation. The absence of iC3b immunoreactivity after EDTA treatment therefore shows that the iC3b detected in Aβ alone and Aβ IC conditions (lanes 1 and 3 of the iC3b blot) was specifically generated by Aβ and Aβ IC complement activation, and not by endogenous levels of iC3b. Compared to Aβ alone, iC3b immunoreactivity was markedly enhanced by Aβ ICs, consistent with enhanced complement activation by Aβ ICs in our earlier experiments. A parallel Western blot for Aβ (6E10 detection antibody) using the Aβ solution employed in the experiment is shown in the left-most lane. Note that bands for iC3b and Aβ aggregate forms typically coincide, as would be predicted given that iC3b is covalently bound to Aβ aggregate forms that can activate complement. B) Densitometry of the blot in panel A, as well as a second blot, prepared in the same way but pre-depleted of endogenous Igs and iC3b, gave a similar pattern of results that did not differ significantly from blot to blot (P = 0.307), with both showing highly-enriched iC3b immunoreactivity in Aβ IC compared to Aβ alone samples (F = 71.65, P = 0.001). For example, integrated optical density of the molecular weight bands designated by the four circled numbers on panel A were increased by 3-fold to 5-fold in the Aβ IC condition.
    Edta, supplied by Amresco, used in various techniques. Bioz Stars score: 99/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/edta/product/Amresco
    Average 99 stars, based on 2 article reviews
    Price from $9.99 to $1999.99
    edta - by Bioz Stars, 2020-03
    99/100 stars
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    SIRT6 promotes the ubiquitination of p27 ( A ) 293 cells were transfected with SIRT6 and vector, twenty four hours after transfection, cells were treated with MG132 or acetyl-leu-leu-norleucinal (ALLN) 5 h before harvesting. Proteins were analyzed by western blot. ( B ) An in vivo ubiquitination assay was performed. 293 cells were transiently transfected with SIRT6 and vector, siRNA-SIRT6 and NC (negative control). Forty two hours later, cells were treated with MG132 for 5 h. Cells were lysed and proceeded for co-immunoprecipitation using the p27 antibody.

    Journal: Aging (Albany NY)

    Article Title: SIRT6 delays cellular senescence by promoting p27Kip1 ubiquitin-proteasome degradation

    doi: 10.18632/aging.101038

    Figure Lengend Snippet: SIRT6 promotes the ubiquitination of p27 ( A ) 293 cells were transfected with SIRT6 and vector, twenty four hours after transfection, cells were treated with MG132 or acetyl-leu-leu-norleucinal (ALLN) 5 h before harvesting. Proteins were analyzed by western blot. ( B ) An in vivo ubiquitination assay was performed. 293 cells were transiently transfected with SIRT6 and vector, siRNA-SIRT6 and NC (negative control). Forty two hours later, cells were treated with MG132 for 5 h. Cells were lysed and proceeded for co-immunoprecipitation using the p27 antibody.

    Article Snippet: Co-immunoprecipitation assays Cells for immunoprecipitation assay were lysed in immunoprecipitation lysis buffer (25 mM Tris-HCl pH 7.4, 150 mM NaCl, 1% Nonidet P-40, 1 mM EDTA, 5% glycerol) containing Protease Inhibitor Mixture (Amresco).

    Techniques: Transfection, Plasmid Preparation, Western Blot, In Vivo, Ubiquitin Assay, Negative Control, Immunoprecipitation

    PIASxα and PTEN interact with each other both in vivo and in vitro . A , co-immunoprecipitation of PIASxα with PTEN from HeLa cells. Cells were transfected with FLAG-PIASxα. 48 h after transfection, cells were harvested and lysed.

    Journal: The Journal of Biological Chemistry

    Article Title: PIASxα Ligase Enhances SUMO1 Modification of PTEN Protein as a SUMO E3 Ligase *

    doi: 10.1074/jbc.M113.508515

    Figure Lengend Snippet: PIASxα and PTEN interact with each other both in vivo and in vitro . A , co-immunoprecipitation of PIASxα with PTEN from HeLa cells. Cells were transfected with FLAG-PIASxα. 48 h after transfection, cells were harvested and lysed.

    Article Snippet: Cells for immunoprecipitation assay were lysed in immunoprecipitation lysis buffer (25 m m Tris-HCl pH 7.4, 150 m m NaCl, 1% Nonidet P-40, 1 m m EDTA, 5% glycerol) containing Protease Inhibitor Mixture (Amresco).

    Techniques: In Vivo, In Vitro, Immunoprecipitation, Transfection

    Complement opsonization of Aβ and Aβ/4G8 ICs A) After exposure to NHS to permit complement activation, Aβ alone and Aβ IC solutions were run on SDS reducing/denaturing Western blots to detect the complement opsonin iC3b. EDTA treatment (lanes 2 and 4 of the iC3b blot) abolished complement activation. The absence of iC3b immunoreactivity after EDTA treatment therefore shows that the iC3b detected in Aβ alone and Aβ IC conditions (lanes 1 and 3 of the iC3b blot) was specifically generated by Aβ and Aβ IC complement activation, and not by endogenous levels of iC3b. Compared to Aβ alone, iC3b immunoreactivity was markedly enhanced by Aβ ICs, consistent with enhanced complement activation by Aβ ICs in our earlier experiments. A parallel Western blot for Aβ (6E10 detection antibody) using the Aβ solution employed in the experiment is shown in the left-most lane. Note that bands for iC3b and Aβ aggregate forms typically coincide, as would be predicted given that iC3b is covalently bound to Aβ aggregate forms that can activate complement. B) Densitometry of the blot in panel A, as well as a second blot, prepared in the same way but pre-depleted of endogenous Igs and iC3b, gave a similar pattern of results that did not differ significantly from blot to blot (P = 0.307), with both showing highly-enriched iC3b immunoreactivity in Aβ IC compared to Aβ alone samples (F = 71.65, P = 0.001). For example, integrated optical density of the molecular weight bands designated by the four circled numbers on panel A were increased by 3-fold to 5-fold in the Aβ IC condition.

    Journal: Alzheimer's & dementia : the journal of the Alzheimer's Association

    Article Title: Peripheral complement interactions with amyloid β peptide (Aβ) in Alzheimer’s disease: 2. Relationship to Aβ immunotherapy

    doi: 10.1016/j.jalz.2017.04.015

    Figure Lengend Snippet: Complement opsonization of Aβ and Aβ/4G8 ICs A) After exposure to NHS to permit complement activation, Aβ alone and Aβ IC solutions were run on SDS reducing/denaturing Western blots to detect the complement opsonin iC3b. EDTA treatment (lanes 2 and 4 of the iC3b blot) abolished complement activation. The absence of iC3b immunoreactivity after EDTA treatment therefore shows that the iC3b detected in Aβ alone and Aβ IC conditions (lanes 1 and 3 of the iC3b blot) was specifically generated by Aβ and Aβ IC complement activation, and not by endogenous levels of iC3b. Compared to Aβ alone, iC3b immunoreactivity was markedly enhanced by Aβ ICs, consistent with enhanced complement activation by Aβ ICs in our earlier experiments. A parallel Western blot for Aβ (6E10 detection antibody) using the Aβ solution employed in the experiment is shown in the left-most lane. Note that bands for iC3b and Aβ aggregate forms typically coincide, as would be predicted given that iC3b is covalently bound to Aβ aggregate forms that can activate complement. B) Densitometry of the blot in panel A, as well as a second blot, prepared in the same way but pre-depleted of endogenous Igs and iC3b, gave a similar pattern of results that did not differ significantly from blot to blot (P = 0.307), with both showing highly-enriched iC3b immunoreactivity in Aβ IC compared to Aβ alone samples (F = 71.65, P = 0.001). For example, integrated optical density of the molecular weight bands designated by the four circled numbers on panel A were increased by 3-fold to 5-fold in the Aβ IC condition.

    Article Snippet: NHS plus 10 mM EDTA (Amresco, Solon, OH) (final concentration in the serum) was employed as a control.

    Techniques: Activation Assay, Western Blot, Generated, Molecular Weight

    Assessments of solvent tolerance and chelator challenge. Potential complications of screening conditions were addressed by challenging Obg GC with various concentrations of DMSO (A) or multiple known chelators (B) followed by measurement of GTPase Obg GC activity using free phosphate quantitation with BIOMOL Green. Protein activity was similar in the presence of 0–5% DMSO and in the presence of three common chelators EDTA, EGTA, and NTA tested 40 μM final concentration. CA significantly decreased the amount of phosphate detected, but it also decreased the background phosphate in the reaction lacking Obg GC (not shown), and was therefore altering the absorbance of the reagent, not acting as a chelator. Asterisk denotes significant difference (ANOVA, P

    Journal: PLoS ONE

    Article Title: Targeting an Essential GTPase Obg for the Development of Broad-Spectrum Antibiotics

    doi: 10.1371/journal.pone.0148222

    Figure Lengend Snippet: Assessments of solvent tolerance and chelator challenge. Potential complications of screening conditions were addressed by challenging Obg GC with various concentrations of DMSO (A) or multiple known chelators (B) followed by measurement of GTPase Obg GC activity using free phosphate quantitation with BIOMOL Green. Protein activity was similar in the presence of 0–5% DMSO and in the presence of three common chelators EDTA, EGTA, and NTA tested 40 μM final concentration. CA significantly decreased the amount of phosphate detected, but it also decreased the background phosphate in the reaction lacking Obg GC (not shown), and was therefore altering the absorbance of the reagent, not acting as a chelator. Asterisk denotes significant difference (ANOVA, P

    Article Snippet: The effect of compounds that may act as Mg2+ chelators and disrupt or inhibit the ObgGC GTPase activity was tested using common chelators including EDTA (Amresco, Solon, OH), ethylene glycol tetra acetic acid (EGTA; Alfa Aesar, Ward Hill, MA), Nitrilotriacetic acid (NTA; TCI, Portland, OR), and citric acid (CA; Sigma Aldrich, St. Louis, MO) at the test-compound concentration used in HTS (40 μM).

    Techniques: Activity Assay, Quantitation Assay, Concentration Assay