Journal: PLoS ONE

Article Title: Studying the Salt Dependence of the Binding of ?70 and ?32 to Core RNA Polymerase Using Luminescence Resonance Energy Transfer

doi: 10.1371/journal.pone.0006490

Figure Lengend Snippet: The effect of common buffer components on the LRET assay. Common buffer components were tested for their ability to alter the interaction 10 nM Tb-core with 20 nM F-σ70 after a 1-hour incubation at 22°C. The average value of A 520 /A 490 for three replicates was determined and normalized to a no-treatment control with a buffer containing 50 mM Tris-HCl pH 7.9, 5% glycerol, and 100 mM NaCl. The error bars represent the normalized standard deviation for a sample size of 4. A) Common solvents (DMSO, ethanol, and methanol) were tested in a range of 0–50% (v/v). B) The non-ionic detergents TritonX-100 and Tween-20 were tested from 0–5% (v/v). C) Glycerol was tested from 2.5–50% (v/v). The minimum glycerol level (2.5%) was due to the glycerol in the buffer of the proteins. D) Bovine serum albumin (BSA) and MgCl 2 were tested from 0–1000 µg/mL and 0–500 mM, respectively.

Article Snippet: Non-ionic detergents such as TritonX-100 and Tween-20 had no effect up to 2.5% ( ).

Techniques: Incubation, Standard Deviation

Journal: PLoS ONE

Article Title: Construction of DNA Hemicatenanes from Two Small Circular DNA Molecules

doi: 10.1371/journal.pone.0119368

Figure Lengend Snippet: Effect of Triton X-100 on the migration of single stranded DNA in polyacrylamide gels. The gel illustrates the smearing of single stranded DNA when loaded on polyacrylamide gels in the presence of Triton X-100. The DNA sample was the same in all lanes and contained a labeled single stranded circle of 235 nucleotides. Two minor bands that show no smearing most probably correspond to contaminating double stranded DNA, as smearing was always observed with many different single strands. Three different brands of Triton-X100 denoted T1–T3 were tested at a final concentration of 0.05%: T1 and T2 were ordinary grade, T3 was highly purified, oxidant-free Triton-X100, sold in glass ampules sealed under nitrogen (Pierce). Lane 1: control, no Triton-X100 added; lane 2: deionized T1; lane 3: T1, untreated; lane 4: T1, heat-treated; lane 5: T1, SDS added at a concentration of 0.005%; lane 6: T3, untreated; lane 7: T1, washed with a 5M NaCl solution; lane 8: T2, untreated. Note that addition of SDS to the samples before loading the gel was a very efficient way to suppress smearing (lane 5).

Article Snippet: Deionization of Triton-X100 with mixed-bed ion exchange resin, or use of highly-purified Triton-X100 sold in glass ampules sealed under nitrogen (Pierce), did not significantly change the result ( ).

Techniques: Migration, Labeling, Concentration Assay, Purification

Journal: Molecular Cancer

Article Title: TGF?1 activates c-Jun and Erk1 via ?V?6 integrin

doi: 10.1186/1476-4598-2-33

Figure Lengend Snippet: Erk1 is associated with focal adhesions. BxPC-3 cells were stimulated with 10 nM of mature TGFβ 1 for ten minutes followed by preparation of the Triton-X100 nonsoluble fraction and precipitation with α V β 6 integrin antibodies. The precipitate was then re-precipitated with anti-FAK antibodies (sc-1688) and analyzed with an Erk1 antibody (sc-93). In part, the cells were preincubated with a TGFβ 1 antibody [ 10 ].

Article Snippet: After preparation of the cytoskeletal fraction by Triton-X100 extraction [ , , ], slides were stained using Alexa Fluor 680 goat anti-mouse IgG (Molecular Probes, Eugene, OR) for Actin, Alexa Fluor 488 donkey anti-rabbit IgG (Molecular Probes) for αV β6 integrin, and Alexa Fluor 350 donkey anti-rabbit IgG (Molecular Probes) for TGFβ1 labeling [ ].

Techniques:

Journal: Molecular Cancer

Article Title: TGF?1 activates c-Jun and Erk1 via ?V?6 integrin

doi: 10.1186/1476-4598-2-33

Figure Lengend Snippet: Paxillin and c-Jun are associated with focal adhesions. BxPC-3 cells were stimulated with 10 nM of mature TGFβ 1 for ten minutes followed by preparation of the Triton-X100 nonsoluble fraction and precipitation with α V β 6 integrin antibodies. The precipitate was then re-precipitated with anti-FAK antibodies (sc-1688) and analyzed with antibodies against p-Paxillin Y118 (2541, Cell Signaling) and p-c-Jun S63 (9621, Cell Signaling). In part, the cells were preincubated with a TGFβ-RII antibody (Santa Cruz), BAPTA-AM and Cytochalasin D, respectively [ 10 ].

Article Snippet: After preparation of the cytoskeletal fraction by Triton-X100 extraction [ , , ], slides were stained using Alexa Fluor 680 goat anti-mouse IgG (Molecular Probes, Eugene, OR) for Actin, Alexa Fluor 488 donkey anti-rabbit IgG (Molecular Probes) for αV β6 integrin, and Alexa Fluor 350 donkey anti-rabbit IgG (Molecular Probes) for TGFβ1 labeling [ ].

Techniques:

Journal: Molecular Cancer

Article Title: TGF?1 activates c-Jun and Erk1 via ?V?6 integrin

doi: 10.1186/1476-4598-2-33

Figure Lengend Snippet: MEKK1 is associated with focal adhesions. BxPC-3 cells were stimulated with 10 nM of mature TGFβ 1 for ten minutes followed by preparation of the Triton-X100 nonsoluble fraction and precipitation with α V β 6 integrin antibodies. The precipitate was then re-precipitated with anti-FAK antibodies (sc-1688) and analyzed with a MEKK1 antibody (sc-448). In part, the cells were preincubated with a TGFβ 1 antibody [ 10 ].

Article Snippet: After preparation of the cytoskeletal fraction by Triton-X100 extraction [ , , ], slides were stained using Alexa Fluor 680 goat anti-mouse IgG (Molecular Probes, Eugene, OR) for Actin, Alexa Fluor 488 donkey anti-rabbit IgG (Molecular Probes) for αV β6 integrin, and Alexa Fluor 350 donkey anti-rabbit IgG (Molecular Probes) for TGFβ1 labeling [ ].

Techniques:

Journal: Molecular Cancer

Article Title: TGF?1 activates c-Jun and Erk1 via ?V?6 integrin

doi: 10.1186/1476-4598-2-33

Figure Lengend Snippet: Pak1 is associated with focal adhesions. BxPC-3 cells were stimulated with 10 nM of mature TGFβ 1 for ten minutes followed by preparation of the Triton-X100 nonsoluble fraction and precipitation with α V β 6 integrin antibodies. The precipitate was then reprecipitated with anti-FAK antibodies (sc-1688), followed by re-precipitation with anti-phospho S (ab9334, Abcam), anti-phospho Thr (ab2286), and analyzed with a Pak1 antibody (71-9300, Zymed). In part, the cells were preincubated with 50 μM PP2 (Calbiochem), and 50 μM PD98059 (Calbiochem), respectively [ 10 ].

Article Snippet: After preparation of the cytoskeletal fraction by Triton-X100 extraction [ , , ], slides were stained using Alexa Fluor 680 goat anti-mouse IgG (Molecular Probes, Eugene, OR) for Actin, Alexa Fluor 488 donkey anti-rabbit IgG (Molecular Probes) for αV β6 integrin, and Alexa Fluor 350 donkey anti-rabbit IgG (Molecular Probes) for TGFβ1 labeling [ ].

Techniques:

Journal: Molecular Cancer

Article Title: TGF?1 activates c-Jun and Erk1 via ?V?6 integrin

doi: 10.1186/1476-4598-2-33

Figure Lengend Snippet: Colocalization of TGFβ 1 , α V β 6 integrin and the cytoskeleton. 10,000 cells(SW48, DLD1, HeLa, keratinocytes) were cultured on glass coverslips in DMEM supplemented with 17% of heat inactivated fetal bovine serum and stimulated with 10 nM of mature TGFβ 1 (from R D Systems) for ten minutes. After preparation of the cytoskeletal fraction by Triton-X100 extraction [ 10 ], slides were stained using Alexa Fluor 680 goat anti-mouse IgG (Molecular Probes, Eugene, OR) for actin (sc-8432, Santa Cruz), Alexa Fluor 488 donkey anti-rabbit IgG (Molecular Probes) for α V β 6 integrin (sc-6617 and sc-6632), and Alexa Fluor 350 donkey anti-rabbit IgG (Molecular Probes) for TGFβ 1 (sc-146) labeling and viewed using a Zeiss LSM-510 confocal microscope [ 10 ]. Magnification 1000 ×.

Article Snippet: After preparation of the cytoskeletal fraction by Triton-X100 extraction [ , , ], slides were stained using Alexa Fluor 680 goat anti-mouse IgG (Molecular Probes, Eugene, OR) for Actin, Alexa Fluor 488 donkey anti-rabbit IgG (Molecular Probes) for αV β6 integrin, and Alexa Fluor 350 donkey anti-rabbit IgG (Molecular Probes) for TGFβ1 labeling [ ].

Techniques: Cell Culture, Staining, Labeling, Microscopy

Journal: Journal of biomedical optics

Article Title: Delivery of Optical Contrast Agents using Triton-X100, Part 1: Reversible permeabilization of live cells for intracellular labeling

doi: 10.1117/1.3090448

Figure Lengend Snippet: Permeabilization of 1483 cell monolayers treated with different concentrations of Triton-X100, as measured by MTT reduction and 3kDa rhodamine-dextran uptake. Cells were treated with Triton-X100 for 10 minutes, incubated in media for 4 hours, and then

Article Snippet: Briefly, subconfluent monolayers of cells (plated 24 hours in advance at 5 × 104 cells/cm2 ) were washed once with cold phosphate buffered saline (PBS; Sigma-Aldrich, St. Louis, MO) and treated with Triton-X100 (Sigma-Aldrich) for 10 minutes at 4°C.

Techniques: MTT Assay, Incubation

Journal: Journal of biomedical optics

Article Title: Delivery of Optical Contrast Agents using Triton-X100, Part 1: Reversible permeabilization of live cells for intracellular labeling

doi: 10.1117/1.3090448

Figure Lengend Snippet: Macromolecule penetration into the cytoplasm and nucleus as a function of Triton-X100 concentration. Subconfluent monolayers of 1483 cells were pre-treated with different concentrations of Triton-X100 for 10 minutes, and then probed with a 1:1:1 mixture

Article Snippet: Briefly, subconfluent monolayers of cells (plated 24 hours in advance at 5 × 104 cells/cm2 ) were washed once with cold phosphate buffered saline (PBS; Sigma-Aldrich, St. Louis, MO) and treated with Triton-X100 (Sigma-Aldrich) for 10 minutes at 4°C.

Techniques: Concentration Assay

Journal: Journal of biomedical optics

Article Title: Delivery of Optical Contrast Agents using Triton-X100, Part 1: Reversible permeabilization of live cells for intracellular labeling

doi: 10.1117/1.3090448

Figure Lengend Snippet: Cumulative macromolecule penetration as a function of time following the administration of 0.55pmol/cell Triton-X100. (a) The time-course of AlexaFluor647 IgG accumulation in the cytoplasm and nucleus of live 1483 cells, averaged over 5 trials. Cells

Article Snippet: Briefly, subconfluent monolayers of cells (plated 24 hours in advance at 5 × 104 cells/cm2 ) were washed once with cold phosphate buffered saline (PBS; Sigma-Aldrich, St. Louis, MO) and treated with Triton-X100 (Sigma-Aldrich) for 10 minutes at 4°C.

Techniques:

Journal: Journal of biomedical optics

Article Title: Delivery of Optical Contrast Agents using Triton-X100, Part 1: Reversible permeabilization of live cells for intracellular labeling

doi: 10.1117/1.3090448

Figure Lengend Snippet: Cell recovery after treatment with Triton-X100. (a) Metabolic activity of 1483 cells 24 hours after treatment with different concentrations of Triton-X100, as measured by the MTT assay. A dose-dependent decrease in the metabolic activity (◊) was

Article Snippet: Briefly, subconfluent monolayers of cells (plated 24 hours in advance at 5 × 104 cells/cm2 ) were washed once with cold phosphate buffered saline (PBS; Sigma-Aldrich, St. Louis, MO) and treated with Triton-X100 (Sigma-Aldrich) for 10 minutes at 4°C.

Techniques: Activity Assay, MTT Assay

Journal: Journal of biomedical optics

Article Title: Delivery of Optical Contrast Agents using Triton-X100, Part 1: Reversible permeabilization of live cells for intracellular labeling

doi: 10.1117/1.3090448

Figure Lengend Snippet: Detection of intra-nuclear markers by immunofluorescence following Triton-X100 treatment. DIC and confocal fluorescence images of 1483 cells labeled with the PC563-hTERT antibody (red) and NCL-hTERT antibody (green) or IgG control antibodies (left panels).

Article Snippet: Briefly, subconfluent monolayers of cells (plated 24 hours in advance at 5 × 104 cells/cm2 ) were washed once with cold phosphate buffered saline (PBS; Sigma-Aldrich, St. Louis, MO) and treated with Triton-X100 (Sigma-Aldrich) for 10 minutes at 4°C.

Techniques: Immunofluorescence, Fluorescence, Labeling

Journal: International Journal of Nanomedicine

Article Title: Cellular entry of nanoparticles via serum sensitive clathrin-mediated endocytosis, and plasma membrane permeabilization

doi: 10.2147/IJN.S29334

Figure Lengend Snippet: Incubation of HeLa cells in the presence of nanoparticles at 4°C in serum-free media resulted in significantly higher Sytox ® Green (Invitrogen Life Technologies, Carlsbad, CA) fluorescence than control cells. Representative images of a HeLa cell incubated with ( A ) Sytox Green, ( B ) nanoparticles and Sytox Green, and ( C ) Triton X100 and Sytox Green in serum-free media at 4°C. The average Sytox Green fluorescence per cell for control, nanoparticle-incubated, and detergent-exposed HeLa cells in ( D ) serum-free media and ( E ) serum-containing media. Abbreviations: NPs, nanoparticles; SFM, serum-free media.

Article Snippet: Alternatively, the fixed cells were incubated for 5 minutes in 0.1% Triton X100 solution (Sigma–Aldrich Corporation, St Louis, MO) and then incubated for 5 minutes in a 0.02% Sytox Green solution (Invitrogen, Grand Island, NY) before mounting.

Techniques: Incubation, Fluorescence

Journal: Autophagy

Article Title: GBA deficiency promotes SNCA/α-synuclein accumulation through autophagic inhibition by inactivated PPP2A

doi: 10.1080/15548627.2015.1086055

Figure Lengend Snippet: SNCA oligomerization and aggregation by loss of GBA function. ( A, B ) The accumulation of high-molecular-weight SNCA species were significantly increased in the TritonX-100 insoluble fraction in LV-GFP-sh Gba #6-infected neurons, and this effect was abolished

Article Snippet: Primary rat cortical neurons were resuspended in 5 volumes of ice-cold TritonX-100 lysis buffer (50 mM Tris, pH 7.4, 175 mM NaCl, 5 mM EDTA, 1% TritonX-100 [Sigma, V900502], 1 mM PMSF), and incubated on ice for 30 min.

Techniques: Molecular Weight, Infection

Journal: The Journal of Neuroscience

Article Title: Membrane Lipid Rafts Are Necessary for the Maintenance of the α7 Nicotinic Acetylcholine Receptor in Somatic Spines of Ciliary Neurons

doi: 10.1523/JNEUROSCI.21-02-00504.2001

Figure Lengend Snippet: α7nAChR is associated with detergent-resistant membrane lipid microdomains. A membrane fraction from St 40–42 CGs was prepared by gradient centrifugation, incubated with Triton X-100 on ice for 30 min, and subjected to discontinuous gradient centrifugation (35–30-0% Optiprep) at 285,000 × g for 4 hr at 4°C. Six fractions were collected from the top ( fraction 1 ) to the bottom ( fraction 6 ). α7nAChR was isolated from each sample with αBTX-conjugated Actigel, separated on 10% SDS-PAGE, and analyzed by Western blotting with a monoclonal antibody against the α7nAChR. An aliquot of each fraction was also analyzed by Western blotting with a polyclonal antibody against caveolin. The Western blots indicate that the bulk of α7nAChR floats to the lower density fractions ( fraction 1, 2 ), indicating its association with detergent-resistant membrane structures. The accumulation of the integral lipid raft protein caveolin in the first two fractions confirms the migration of the detergent-insoluble components of the membrane to the low-density portion of the gradient.

Article Snippet: The samples were then washed three times with solubilization solution, twice with of 0.1 m phosphate buffer, pH 7.3, 0.5% Triton X-100, and 1 m NaCl, and twice with 0.1 m phosphate buffer, pH 7.3, and 0.5% Triton X-100 and resuspended in 50 μl of Laemmli sample buffer.

Techniques: Gradient Centrifugation, Incubation, Isolation, SDS Page, Western Blot, Migration

Journal: Interdisciplinary Toxicology

Article Title: Damascenine induced hepatotoxicity and nephrotoxicity in mice and in vitro assessed human erythrocyte toxicity

doi: 10.1515/intox-2015-0018

Figure Lengend Snippet: Photomicrographs (G×40) showing morphology of erythrocytes in the absence and presence of damascenine: (A) positive control cells incubated with 1% Triton-X100; (B) vehicle cells incubated with 10% DMSO; (C) cells incubated with 250 μg/ml of DA; (D) cells incubated with 1000 μg/ml of DA.

Article Snippet: Chemicals and reagents Triton-X100, dimethyl sulfoxide (DMSO), petroleum ether (bp 30–40 °C), ammonium hydroxide solution (28% NH3 in H2 O) and hydrochloric acid (37%) were obtained from Sigma-Aldrich Chemical (St. Louis, MO, USA).

Techniques: Positive Control, Incubation

Journal: MethodsX

Article Title: Colorimetric quantification of sucrose in presence of thermo-sensitive polymers present in aqueous two-phase systems

doi: 10.1016/j.mex.2014.09.006

Figure Lengend Snippet: Standard curves of phenol–sulfuric acid method conducted at an ice-cold temperature (A) and room temperature (B). (A) -×-: sucrose; ⋯+⋯: sucrose + Triton X-114 (0.05%, w/v); ⋯□⋯: sucrose + Triton X-114 (0.5%, w/v); ⋯○⋯: sucrose + Triton X-114 (5%, w/v); (B) -♦-: sucrose; -■-: sucrose + Triton X-114 (0.5%, w/v); -▴- sucrose + Triton X-100 (0.5%, w/v); -●-: sucrose + EO50PO50 (0.5%, w/v)

Article Snippet: Materials • EO50PO50 (Sigma, Cat. No. 438189) • Phenol (Scharlau, Cat. No. FE04821000) • Sucrose (Sigma, Cat. No. S5106) • Sulfuric acid (Merck, Cat. No. 1.00731.1000) • Triton X-100 (Sigma, Cat. No. T9284) • Triton X-114 (Sigma, Cat. No. X144) Ultra-pure water was used throughout the experiment.

Techniques:

Journal: Neurochemical Research

Article Title: Sazetidine-A Activates and Desensitizes Native α7 Nicotinic Acetylcholine Receptors

doi: 10.1007/s11064-014-1302-6

Figure Lengend Snippet: Concentration dependence of sazetidine-A-evoked responses in SH-SY5Y cells. SH-SY5Y cells loaded with fluo-3 AM were stimulated with sazetidine-A (0.1–100 μM) in the presence ( solid black circles ) or absence ( open black circles ) of PNU-120596 (PNU1; 10 μΜ). Fluorescence at 538 nm was measured for 20 s following stimulation with sazetidine-A. The increase in fluorescence at 20 s is presented as a percentage of the maximum fluorescence determined by addition of 0.2 % triton X-100 minus the minimum fluorescence quenched by 350 mM MnCl 2 . Points represent the mean ± SEM from 8 independent experiments. Data were fitted to the Hill equation and EC 50 values for sazatidine-A in the absence and presence of PNU-120596 were calculated to be 4.2 and 0.4 µM respectively

Article Snippet: Materials Triton X-100, (-)-nicotine hydrogen tartrate and mecamylamine hydrochloride, were purchased from Sigma-Aldrich (Poole, Dorset, UK); B27, l -glutamine, antibiotics, fluo-3 AM, fura-2 AM, and pluronic f127 were obtained from Life Technologies (Paisley, UK); sazetidine-A dihydrochloride, tetrodotoxin citrate, methyllycaconitine citrate and 5-iodo-A85380 dihydrochloride were purchased from Tocris Bioscience (Avonmouth, UK); PNU-120596 and PNU-282987 were provided by Pfizer Inc. USA; general reagents were purchased from Fisher Scientific (Loughborough, UK).

Techniques: Concentration Assay, Fluorescence

Journal: Neurochemical Research

Article Title: Sazetidine-A Activates and Desensitizes Native α7 Nicotinic Acetylcholine Receptors

doi: 10.1007/s11064-014-1302-6

Figure Lengend Snippet: Intracellular calcium elevations evoked by sazetidine-A in SH-SY5Y cell populations. SH-SY5Y cells loaded with fluo-3 AM were preincubated for 10 min with or without antagonist (mecamylamine (Mec), 30 μM, chequered bars , or MLA, 100 nM, crosshatched bars ) and/or PNU-120596 (PNU1, 10 μM, grey bars ) before addition of sazetidine-A (Saz, 10 μΜ, a , b ; 100 μM, c , d ) in the presence ( grey bars ) or absence ( white bars ) of PNU-120596 (10 μM). Fluorescence at 538 nm was monitored for 20 s; representative traces are shown ( b , d ; some antagonist curves omitted for clarity). The increase in fluorescence at 20 s is presented as a percentage of the maximum fluorescence determined by addition of 0.2 % triton X-100 minus the minimum fluorescence quenched by 350 mM MnCl 2 ( a , c ). Bars represent the mean ± SEM of at least 4 independent experiments; * P

Article Snippet: Materials Triton X-100, (-)-nicotine hydrogen tartrate and mecamylamine hydrochloride, were purchased from Sigma-Aldrich (Poole, Dorset, UK); B27, l -glutamine, antibiotics, fluo-3 AM, fura-2 AM, and pluronic f127 were obtained from Life Technologies (Paisley, UK); sazetidine-A dihydrochloride, tetrodotoxin citrate, methyllycaconitine citrate and 5-iodo-A85380 dihydrochloride were purchased from Tocris Bioscience (Avonmouth, UK); PNU-120596 and PNU-282987 were provided by Pfizer Inc. USA; general reagents were purchased from Fisher Scientific (Loughborough, UK).

Techniques: Fluorescence

Journal: Nanomaterials

Article Title: Effect of Nanotube Film Thickness on the Performance of Nanotube-Silicon Hybrid Solar Cells

doi: 10.3390/nano3040655

Figure Lengend Snippet: UV-vis-NIR spectra of thin vacuum filtration films of single walled carbon nanotubes (SWCNTs) on glass, made from aqueous TritonX-100 suspensions of large diameter arc-discharge material.

Article Snippet: Preparation of SWCNT Suspension Arc-discharge SWCNTs (5 mg, P3-SWCNT, Carbon Solutions Inc., Riverside, CA, USA) were bath sonicated at ~50 WRMS for 1 h in an aqueous solution of TritonX-100 (50 mL, 1% v /v , Sigma, Castle Hill, Australia).

Techniques: Suction Filtration