tris buffer  (Millipore)


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    Name:
    Trometamol
    Description:
    This product is provided as delivered and specified by the issuing Pharmacopoeia All information provided in support of this product including SDS and any product information leaflets have been developed and issued under the Authority of the issuing Pharmacopoeia For further information and support please go to the website of the issuing Pharmacopoeia
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    t2550000
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    Structured Review

    Millipore tris buffer
    Trometamol
    This product is provided as delivered and specified by the issuing Pharmacopoeia All information provided in support of this product including SDS and any product information leaflets have been developed and issued under the Authority of the issuing Pharmacopoeia For further information and support please go to the website of the issuing Pharmacopoeia
    https://www.bioz.com/result/tris buffer/product/Millipore
    Average 99 stars, based on 817 article reviews
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    tris buffer - by Bioz Stars, 2020-07
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    Images

    1) Product Images from "The role of irradiance and C-use strategies in tropical macroalgae photosynthetic response to ocean acidification"

    Article Title: The role of irradiance and C-use strategies in tropical macroalgae photosynthetic response to ocean acidification

    Journal: Scientific Reports

    doi: 10.1038/s41598-018-27333-0

    Inhibition of P gmax in three fleshy ( a ) and five calcifying ( b ) tropical macroalgae as inhibitor block various bicarbonate uptake pathways: external carbonic anhydrase (CA ext ) with acetazolamide (AZ), AE protein by pyridoxal (5) phosphate (PLP), proton pump acidification by Tris buffer and ATPase H + pumps by sodium orthovanadate.
    Figure Legend Snippet: Inhibition of P gmax in three fleshy ( a ) and five calcifying ( b ) tropical macroalgae as inhibitor block various bicarbonate uptake pathways: external carbonic anhydrase (CA ext ) with acetazolamide (AZ), AE protein by pyridoxal (5) phosphate (PLP), proton pump acidification by Tris buffer and ATPase H + pumps by sodium orthovanadate.

    Techniques Used: Inhibition, Blocking Assay, Plasmid Purification

    2) Product Images from "ICP27 Phosphorylation Site Mutants Display Altered Functional Interactions with Cellular Export Factors Aly/REF and TAP/NXF1 but Are Able To Bind Herpes Simplex Virus 1 RNA ▿"

    Article Title: ICP27 Phosphorylation Site Mutants Display Altered Functional Interactions with Cellular Export Factors Aly/REF and TAP/NXF1 but Are Able To Bind Herpes Simplex Virus 1 RNA ▿

    Journal: Journal of Virology

    doi: 10.1128/JVI.01388-09

    Expression of 15 N-labeled ICP27 N terminus and S16,18,114A and S16,18,114E phosphorylation point mutants. Rosetta E. coli cells were transformed with either the pET21b wild-type ICP27 N terminus or phosphorylation point mutant S16,18,114A or S16,18,114E expression plasmids. Cells were grown in Neidhart's minimal media supplemented with 15 NH 4 Cl. Protein expression was induced with IPTG, and 6× His-tagged proteins were purified using Ni-NTA agarose under native protein purification conditions. A portion of the elution fractions from the Ni-NTA column was separated on a 10 to 20% Tris gradient gel, and the gel was stained with Coomassie blue.
    Figure Legend Snippet: Expression of 15 N-labeled ICP27 N terminus and S16,18,114A and S16,18,114E phosphorylation point mutants. Rosetta E. coli cells were transformed with either the pET21b wild-type ICP27 N terminus or phosphorylation point mutant S16,18,114A or S16,18,114E expression plasmids. Cells were grown in Neidhart's minimal media supplemented with 15 NH 4 Cl. Protein expression was induced with IPTG, and 6× His-tagged proteins were purified using Ni-NTA agarose under native protein purification conditions. A portion of the elution fractions from the Ni-NTA column was separated on a 10 to 20% Tris gradient gel, and the gel was stained with Coomassie blue.

    Techniques Used: Expressing, Labeling, Transformation Assay, Mutagenesis, Purification, Protein Purification, Staining

    3) Product Images from "Basic Residues of β-Sheet A Contribute to Heparin Binding and Activation of Vaspin (Serpin A12) *"

    Article Title: Basic Residues of β-Sheet A Contribute to Heparin Binding and Activation of Vaspin (Serpin A12) *

    Journal: The Journal of Biological Chemistry

    doi: 10.1074/jbc.M116.748020

    Secreted vaspin by HaCaT cells is retained in the extracellular matrix. A , vaspin concentrations in supernatants of HaCaT cells transfected with an empty control vector or with human vaspin after incubation with Tris buffer for 1 h. B , HaCaT cells were treated with heparinase for 1 h, NaCl for 10 min, or Triton X-100 for 15 min, and vaspin concentration was measured in the cell supernatants via ELISA. Treatment with heparinase and NaCl resulted in significantly increased vaspin supernatant concentrations. Data are expressed as relative to untreated controls. Presented are the mean ± S.E. ( error bars ) values of at least three experiments performed in triplicates with the exception of the Triton X-100 condition, which was performed twice. Data were analyzed via Kruskal-Wallis test and Dunn's multiple comparisons test; *, p
    Figure Legend Snippet: Secreted vaspin by HaCaT cells is retained in the extracellular matrix. A , vaspin concentrations in supernatants of HaCaT cells transfected with an empty control vector or with human vaspin after incubation with Tris buffer for 1 h. B , HaCaT cells were treated with heparinase for 1 h, NaCl for 10 min, or Triton X-100 for 15 min, and vaspin concentration was measured in the cell supernatants via ELISA. Treatment with heparinase and NaCl resulted in significantly increased vaspin supernatant concentrations. Data are expressed as relative to untreated controls. Presented are the mean ± S.E. ( error bars ) values of at least three experiments performed in triplicates with the exception of the Triton X-100 condition, which was performed twice. Data were analyzed via Kruskal-Wallis test and Dunn's multiple comparisons test; *, p

    Techniques Used: Transfection, Plasmid Preparation, Incubation, Concentration Assay, Enzyme-linked Immunosorbent Assay

    4) Product Images from "Formation and spreading of TDP-43 aggregates in cultured neuronal and glial cells demonstrated by time-lapse imaging"

    Article Title: Formation and spreading of TDP-43 aggregates in cultured neuronal and glial cells demonstrated by time-lapse imaging

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0179375

    Adenovirus-transduced artificial TDP-43 cytoplasmic aggregates have insoluble properties against various detergents. ( a, b ) Sarkosyl-fractionated cell lysates with Tris (T), 1% TritonX-100 (X), 1% sarkosyl (S) and 1 × SDS-sample (P) buffers were examined using phospho (p)-TDP-43 (pSer409/Ser410) ( a ) or C-terminal TDP-43 (405–414) antibody ( b ). ( c, d ) RIPA/urea-fractionated cell lysates with RIPA (R) and urea (U) buffers were examined using pTDP-43 ( c ) or C-terminal TDP-43 (405–414) antibody ( d ). The 74 and 52 kDa bands correspond to phosphorylated DsRed-tagged WT (←pWT) and CTF (←pCTF) TDP-43, respectively ( a, c ). The 72 and 50 kDa bands correspond to non-phosphorylated DsRed-tagged WT (←WT) and CTF (←CTF) TDP-43, respectively. Endogenous rat TDP-43 band at 43 kDa was indicated as arrowhead ( b, d ). ( a-d ) Full length gels and blots are shown.
    Figure Legend Snippet: Adenovirus-transduced artificial TDP-43 cytoplasmic aggregates have insoluble properties against various detergents. ( a, b ) Sarkosyl-fractionated cell lysates with Tris (T), 1% TritonX-100 (X), 1% sarkosyl (S) and 1 × SDS-sample (P) buffers were examined using phospho (p)-TDP-43 (pSer409/Ser410) ( a ) or C-terminal TDP-43 (405–414) antibody ( b ). ( c, d ) RIPA/urea-fractionated cell lysates with RIPA (R) and urea (U) buffers were examined using pTDP-43 ( c ) or C-terminal TDP-43 (405–414) antibody ( d ). The 74 and 52 kDa bands correspond to phosphorylated DsRed-tagged WT (←pWT) and CTF (←pCTF) TDP-43, respectively ( a, c ). The 72 and 50 kDa bands correspond to non-phosphorylated DsRed-tagged WT (←WT) and CTF (←CTF) TDP-43, respectively. Endogenous rat TDP-43 band at 43 kDa was indicated as arrowhead ( b, d ). ( a-d ) Full length gels and blots are shown.

    Techniques Used:

    5) Product Images from "Selective oxidation of B800 bacteriochlorophyll a in photosynthetic light-harvesting protein LH2"

    Article Title: Selective oxidation of B800 bacteriochlorophyll a in photosynthetic light-harvesting protein LH2

    Journal: Scientific Reports

    doi: 10.1038/s41598-019-40082-y

    Electronic absorption spectra of native LH2 (black) and oxidized LH2 (red) in 20 mM Tris-HCl buffer containing 0.1% n- dodecyl- β -D-maltoside (pH = 8.0). Spectra were normalized at Q y peaks of B850 BChl a . Fluorescence emission spectrum of oxidized LH2 by excitation at 680 nm (blue) is shown with its absorption spectrum. Insert shows overlapped absorption spectra of native LH2 and oxidized LH2 in Q y region.
    Figure Legend Snippet: Electronic absorption spectra of native LH2 (black) and oxidized LH2 (red) in 20 mM Tris-HCl buffer containing 0.1% n- dodecyl- β -D-maltoside (pH = 8.0). Spectra were normalized at Q y peaks of B850 BChl a . Fluorescence emission spectrum of oxidized LH2 by excitation at 680 nm (blue) is shown with its absorption spectrum. Insert shows overlapped absorption spectra of native LH2 and oxidized LH2 in Q y region.

    Techniques Used: Fluorescence

    CD spectra of native LH2 (black) and oxidized LH2 (red) in UV (left) and Q y regions (right) in 20 mM Tris-HCl buffer containing 0.1% n- dodecyl- β -D-maltoside (pH = 8.0). Q y absorbance values of B850 BChl a in LH2 samples used for measurements were 0.9.
    Figure Legend Snippet: CD spectra of native LH2 (black) and oxidized LH2 (red) in UV (left) and Q y regions (right) in 20 mM Tris-HCl buffer containing 0.1% n- dodecyl- β -D-maltoside (pH = 8.0). Q y absorbance values of B850 BChl a in LH2 samples used for measurements were 0.9.

    Techniques Used:

    FM-AFM images of native LH2 ( A ) and oxidized LH2 ( B ) adsorbed on mica taken in 20 mM Tris buffer containing 150 mM NaCl (pH 8.0). Left: wide images. Middle: locally enlarged images of single LH2 proteins. Right: overlapped height-profiles of ten proteins.
    Figure Legend Snippet: FM-AFM images of native LH2 ( A ) and oxidized LH2 ( B ) adsorbed on mica taken in 20 mM Tris buffer containing 150 mM NaCl (pH 8.0). Left: wide images. Middle: locally enlarged images of single LH2 proteins. Right: overlapped height-profiles of ten proteins.

    Techniques Used:

    6) Product Images from "Synthesis of site-specific antibody-drug conjugates by ADP-ribosyl cyclases"

    Article Title: Synthesis of site-specific antibody-drug conjugates by ADP-ribosyl cyclases

    Journal: Science Advances

    doi: 10.1126/sciadv.aba6752

    Generation and in vitro evaluation of anti-HER2 ARC-ADC. ( A ) Scheme for generating anti-HER2 ARC-ADC. r.t., room temperature. ( B ) Conjugation kinetics of drug linker to CD38 C-fusion IgG. 2′-Cl-araNAD + –MMAF (1 mM) was incubated with CD38 C-fusion IgG (10 μM) in 50 mM tris buffer (pH 8.5) for various amounts of time on ice. The residual enzymatic activity determined by fluorescence-based activity assays was plotted as a function of incubation time. ( C and D ) Mass spectra of light chains (C) and heavy chains (D) for CD38 C-fusion IgG and anti-HER2 ARC-ADC. MW, molecular weight. ( E ) X-ray structure of human CD38 catalytic domain with 2′-Cl-araNAD + covalently attached to Glu 226 residue. ( F ) Flow cytometric analysis of HER2 expression for four breast cancer cell lines. ( G to J ) In vitro cytotoxicity of anti-HER2 ARC-ADC. HCC1954 (G), MCF7 (H), MDA-MB-231 (I), and MDA-MB-468 (J) cells with varied levels of HER2 expression were incubated for 72 hours at 37°C with 5% CO 2 in the presence of various concentrations of ARC-ADC, 2′-Cl-araNAD + –MMAF, Herceptin, and CD38 C-fusion IgG. Cell viability was measured by MTT assays. Cells treated with culture media and 5 μM paclitaxel were included as 100% viability and 0% viability controls, respectively.
    Figure Legend Snippet: Generation and in vitro evaluation of anti-HER2 ARC-ADC. ( A ) Scheme for generating anti-HER2 ARC-ADC. r.t., room temperature. ( B ) Conjugation kinetics of drug linker to CD38 C-fusion IgG. 2′-Cl-araNAD + –MMAF (1 mM) was incubated with CD38 C-fusion IgG (10 μM) in 50 mM tris buffer (pH 8.5) for various amounts of time on ice. The residual enzymatic activity determined by fluorescence-based activity assays was plotted as a function of incubation time. ( C and D ) Mass spectra of light chains (C) and heavy chains (D) for CD38 C-fusion IgG and anti-HER2 ARC-ADC. MW, molecular weight. ( E ) X-ray structure of human CD38 catalytic domain with 2′-Cl-araNAD + covalently attached to Glu 226 residue. ( F ) Flow cytometric analysis of HER2 expression for four breast cancer cell lines. ( G to J ) In vitro cytotoxicity of anti-HER2 ARC-ADC. HCC1954 (G), MCF7 (H), MDA-MB-231 (I), and MDA-MB-468 (J) cells with varied levels of HER2 expression were incubated for 72 hours at 37°C with 5% CO 2 in the presence of various concentrations of ARC-ADC, 2′-Cl-araNAD + –MMAF, Herceptin, and CD38 C-fusion IgG. Cell viability was measured by MTT assays. Cells treated with culture media and 5 μM paclitaxel were included as 100% viability and 0% viability controls, respectively.

    Techniques Used: In Vitro, Conjugation Assay, Incubation, Activity Assay, Fluorescence, Molecular Weight, Expressing, Multiple Displacement Amplification, MTT Assay

    7) Product Images from "Basic Residues of β-Sheet A Contribute to Heparin Binding and Activation of Vaspin (Serpin A12) *"

    Article Title: Basic Residues of β-Sheet A Contribute to Heparin Binding and Activation of Vaspin (Serpin A12) *

    Journal: The Journal of Biological Chemistry

    doi: 10.1074/jbc.M116.748020

    Secreted vaspin by HaCaT cells is retained in the extracellular matrix. A , vaspin concentrations in supernatants of HaCaT cells transfected with an empty control vector or with human vaspin after incubation with Tris buffer for 1 h. B , HaCaT cells were treated with heparinase for 1 h, NaCl for 10 min, or Triton X-100 for 15 min, and vaspin concentration was measured in the cell supernatants via ELISA. Treatment with heparinase and NaCl resulted in significantly increased vaspin supernatant concentrations. Data are expressed as relative to untreated controls. Presented are the mean ± S.E. ( error bars ) values of at least three experiments performed in triplicates with the exception of the Triton X-100 condition, which was performed twice. Data were analyzed via Kruskal-Wallis test and Dunn's multiple comparisons test; *, p
    Figure Legend Snippet: Secreted vaspin by HaCaT cells is retained in the extracellular matrix. A , vaspin concentrations in supernatants of HaCaT cells transfected with an empty control vector or with human vaspin after incubation with Tris buffer for 1 h. B , HaCaT cells were treated with heparinase for 1 h, NaCl for 10 min, or Triton X-100 for 15 min, and vaspin concentration was measured in the cell supernatants via ELISA. Treatment with heparinase and NaCl resulted in significantly increased vaspin supernatant concentrations. Data are expressed as relative to untreated controls. Presented are the mean ± S.E. ( error bars ) values of at least three experiments performed in triplicates with the exception of the Triton X-100 condition, which was performed twice. Data were analyzed via Kruskal-Wallis test and Dunn's multiple comparisons test; *, p

    Techniques Used: Transfection, Plasmid Preparation, Incubation, Concentration Assay, Enzyme-linked Immunosorbent Assay

    8) Product Images from "Succinyl-CoA synthetase (SUCLA2) deficiency in two siblings with impaired activity of other mitochondrial oxidative enzymes in skeletal muscle without mitochondrial DNA depletion"

    Article Title: Succinyl-CoA synthetase (SUCLA2) deficiency in two siblings with impaired activity of other mitochondrial oxidative enzymes in skeletal muscle without mitochondrial DNA depletion

    Journal: Molecular genetics and metabolism

    doi: 10.1016/j.ymgme.2016.11.005

    A-SCS activity measurement by LC-MS/MS. (A) Time dependency was measured in control line and patient line at 37°C for 0, 3, 6, 9 and 12 min with 50 mM Tris buffer pH 8.0, 5 mM MgCl 2 , 10 mM D4-succinate, 1 mM ATP, 1 mM CoA, oligomycin 2 µg/ml and homogenate protein concentration 1mg/ml. Activity linearity is shown within 9min. (B) Protein dependency was measured in control line for 6 min at different protein concentrations, 0, 0.5, 0.75, 1 and 1.25 mg/ml. Other conditions were same as (A). Linear for protein concentrations
    Figure Legend Snippet: A-SCS activity measurement by LC-MS/MS. (A) Time dependency was measured in control line and patient line at 37°C for 0, 3, 6, 9 and 12 min with 50 mM Tris buffer pH 8.0, 5 mM MgCl 2 , 10 mM D4-succinate, 1 mM ATP, 1 mM CoA, oligomycin 2 µg/ml and homogenate protein concentration 1mg/ml. Activity linearity is shown within 9min. (B) Protein dependency was measured in control line for 6 min at different protein concentrations, 0, 0.5, 0.75, 1 and 1.25 mg/ml. Other conditions were same as (A). Linear for protein concentrations

    Techniques Used: Activity Assay, Liquid Chromatography with Mass Spectroscopy, Mass Spectrometry, Protein Concentration

    9) Product Images from "Pleiotropic effects of the vacuolar ABC transporter MLT1 of Candida albicans on cell function and virulence"

    Article Title: Pleiotropic effects of the vacuolar ABC transporter MLT1 of Candida albicans on cell function and virulence

    Journal: Biochemical Journal

    doi: 10.1042/BCJ20160024

    Mlt1p levels are required to maintain growth in the presence of NiSO 4 , methotrexate and for accumulation of NBD-PC into the vacuolar lumen ( A ) Comparison of growth by spot dilution assays of C. albicans WT (SC5314), mlt1 Δ/Δ and mlt1 Δ/Δ ::MLT1 cells. A 5-fold serial dilution of each strain was spotted on to NiSO 4 and MTX at the indicated concentrations, in YEPD agar plates and grown for 48 h at 30°C. ( B ) Deletion of MLT1 results in the loss of accumulation of NBD-PC in the vacuolar lumen. C. albicans WT (SC5314), mlt1 Δ/Δ and mlt1 Δ/Δ ::MLT1 cells were grown to mid-exponential phase and incubated with NBD-PC. After incubation, cells were washed and stained with FM4-64. Samples were prepared for microscopy and photographed under a confocal microscope. Scale bar, 10 μm. ( C ) NBD-PC accumulation into the vacuolar lumen of C. albicans is energy-dependent. Sodium azide treatment was given to mid-exponential-phase WT C. albicans (SC5314) cells before their incubation with NBD-PC. The cells were photographed under a confocal microscope. ( D ) NBD-PC accumulation into the vacuolar lumen is energy-dependent. Vacuoles were isolated from WT and mlt1 Δ/Δ mutant C. albicans using a Ficoll gradient ultracentrifuge-based method. Equal amounts (25 μg) of purified vacuolar vesicles were then incubated at 30°C for 30 min in buffer containing 10 μM NBD-PC and 10 μM FM4-64. A transport assay was carried out under two conditions: one in the absence of energy source ATP and another in the presence of 5 mM ATP. After incubation, samples were washed with ice-cold Tris/sucrose buffer containing 3% fatty-acid-free BSA and observed under a confocal microscope. In the presence of ATP, the isolated vacuoles from WT C. albicans showed NBD-PC accumulation. There was no NBD-PC accumulation in the absence of ATP in WT cells and in vacuoles isolated from mlt1 Δ/Δ mutant. DIC, differential interference contrast.
    Figure Legend Snippet: Mlt1p levels are required to maintain growth in the presence of NiSO 4 , methotrexate and for accumulation of NBD-PC into the vacuolar lumen ( A ) Comparison of growth by spot dilution assays of C. albicans WT (SC5314), mlt1 Δ/Δ and mlt1 Δ/Δ ::MLT1 cells. A 5-fold serial dilution of each strain was spotted on to NiSO 4 and MTX at the indicated concentrations, in YEPD agar plates and grown for 48 h at 30°C. ( B ) Deletion of MLT1 results in the loss of accumulation of NBD-PC in the vacuolar lumen. C. albicans WT (SC5314), mlt1 Δ/Δ and mlt1 Δ/Δ ::MLT1 cells were grown to mid-exponential phase and incubated with NBD-PC. After incubation, cells were washed and stained with FM4-64. Samples were prepared for microscopy and photographed under a confocal microscope. Scale bar, 10 μm. ( C ) NBD-PC accumulation into the vacuolar lumen of C. albicans is energy-dependent. Sodium azide treatment was given to mid-exponential-phase WT C. albicans (SC5314) cells before their incubation with NBD-PC. The cells were photographed under a confocal microscope. ( D ) NBD-PC accumulation into the vacuolar lumen is energy-dependent. Vacuoles were isolated from WT and mlt1 Δ/Δ mutant C. albicans using a Ficoll gradient ultracentrifuge-based method. Equal amounts (25 μg) of purified vacuolar vesicles were then incubated at 30°C for 30 min in buffer containing 10 μM NBD-PC and 10 μM FM4-64. A transport assay was carried out under two conditions: one in the absence of energy source ATP and another in the presence of 5 mM ATP. After incubation, samples were washed with ice-cold Tris/sucrose buffer containing 3% fatty-acid-free BSA and observed under a confocal microscope. In the presence of ATP, the isolated vacuoles from WT C. albicans showed NBD-PC accumulation. There was no NBD-PC accumulation in the absence of ATP in WT cells and in vacuoles isolated from mlt1 Δ/Δ mutant. DIC, differential interference contrast.

    Techniques Used: Serial Dilution, Incubation, Staining, Microscopy, Isolation, Mutagenesis, Purification, Transport Assay

    10) Product Images from "Purification and Characterization of Put1p from Saccharomyces cerevisiae"

    Article Title: Purification and Characterization of Put1p from Saccharomyces cerevisiae

    Journal: Archives of biochemistry and biophysics

    doi: 10.1016/j.abb.2010.04.020

    UV-visible spectrum of purified Put1p (C-terminal 6xHis tagged form) in 20 mM Tris-HCl buffer (pH 8.0) containing 100 mM NaCl and 0.25% octyl-glucoside. Concentrations of total and flavin-bound Put1p protein are 16.1 µM and 7 µM, respectively.
    Figure Legend Snippet: UV-visible spectrum of purified Put1p (C-terminal 6xHis tagged form) in 20 mM Tris-HCl buffer (pH 8.0) containing 100 mM NaCl and 0.25% octyl-glucoside. Concentrations of total and flavin-bound Put1p protein are 16.1 µM and 7 µM, respectively.

    Techniques Used: Purification

    11) Product Images from "Study of the Assembly of Vesicular Stomatitis Virus N Protein: Role of the P Protein"

    Article Title: Study of the Assembly of Vesicular Stomatitis Virus N Protein: Role of the P Protein

    Journal: Journal of Virology

    doi:

    Gel filtration profiles of the N/P protein complexes at pH 7.5 (A), 6.0 (C), 5.0 (E), and 4.0 (G). The N and P proteins were coexpressed in E. coli harboring plasmid pET-N/P. Following protein expression, the N/P protein complexes were isolated by Ni affinity chromatography. These partially purified protein complexes were dialyzed in either 0.05 M Tris (pH 7.5) containing 300 mM NaCl or 0.1 M citrate (pH 6.0, 5.0, or 4.0) containing 0.25 M NaCl. Following dialysis, the complexes were chromatographed on a Sephacryl S-300 size exclusion column (16/60; Pharmacia) at a flow rate of 1 ml/min. In each case, the elution buffer was the same as the dialysis buffer. The horizontal axis shows the elution time in minutes; the vertical axis shows A 280 . The proteins contained in the peaks from each purification were analyzed by SDS-polyacrylamide gel electrophoresis (gel B for panel A, gel D for panel C, gel F for panel E, and gel H for panel G). (B, D, F, and H). The proteins were electrophoresed on 10% gels and stained with Coomassie brilliant blue R-250. Lane numbers correspond to the peak numbers on the corresponding chromatogram to the left. Lanes containing molecular weight standards are labeled MW; positions are indicated in kilodaltons.
    Figure Legend Snippet: Gel filtration profiles of the N/P protein complexes at pH 7.5 (A), 6.0 (C), 5.0 (E), and 4.0 (G). The N and P proteins were coexpressed in E. coli harboring plasmid pET-N/P. Following protein expression, the N/P protein complexes were isolated by Ni affinity chromatography. These partially purified protein complexes were dialyzed in either 0.05 M Tris (pH 7.5) containing 300 mM NaCl or 0.1 M citrate (pH 6.0, 5.0, or 4.0) containing 0.25 M NaCl. Following dialysis, the complexes were chromatographed on a Sephacryl S-300 size exclusion column (16/60; Pharmacia) at a flow rate of 1 ml/min. In each case, the elution buffer was the same as the dialysis buffer. The horizontal axis shows the elution time in minutes; the vertical axis shows A 280 . The proteins contained in the peaks from each purification were analyzed by SDS-polyacrylamide gel electrophoresis (gel B for panel A, gel D for panel C, gel F for panel E, and gel H for panel G). (B, D, F, and H). The proteins were electrophoresed on 10% gels and stained with Coomassie brilliant blue R-250. Lane numbers correspond to the peak numbers on the corresponding chromatogram to the left. Lanes containing molecular weight standards are labeled MW; positions are indicated in kilodaltons.

    Techniques Used: Filtration, Plasmid Preparation, Positron Emission Tomography, Expressing, Isolation, Affinity Chromatography, Purification, Flow Cytometry, Polyacrylamide Gel Electrophoresis, Staining, Molecular Weight, Labeling

    12) Product Images from "A Novel Immunoprecipitation Strategy Identifies a Unique Functional Mimic of the Glial Cell Line-Derived Neurotrophic Factor Family Ligands in the Pathogen Trypanosoma cruzi "

    Article Title: A Novel Immunoprecipitation Strategy Identifies a Unique Functional Mimic of the Glial Cell Line-Derived Neurotrophic Factor Family Ligands in the Pathogen Trypanosoma cruzi

    Journal: Infection and Immunity

    doi: 10.1128/IAI.00411-08

    Identification and specificity of TGFL. (A) PcIP. The indicated growth factor receptor-Fc chimeras (GFRα-1, RET, TGF-β-RII, FGF-R1α, TNF-RI, and P75) (each, 0.5 μg) were incubated overnight 4°C with T. cruzi trypomastigote lysate (∼25 to 30 μg), adsorbed to protein G-Sepharose, washed, eluted with 2% SDS, run on a nonreducing SDS-PAGE gel, blotted onto nitrocellulose, probed with chagasic sera followed by HRP-labeled secondary anti-human IgG antibody, and visualized with the ECL system for 1 second and 20 seconds. Numbers on the right represent molecular masses in kDa. The experiment was repeated more than 15 times with nearly identical results. Control lanes, lysate adsorbed to protein G-Sepharose without premixing with receptor ( T. cruzi +) and receptor adsorbed to protein G-Sepharose without premixing with lysate (receptor +); adsorption of lysate-receptor mixtures to protein G-Sepharose is denoted by appropriate + and − signs in the corresponding lanes. (B) Coomassie blue staining of TGFL (1.9 μg, equivalent to 2.3 × 10 8 T. cruzi trypomastigotes, Silvio strain) twice purified on GFRα-1/protein G-Sepharose column after discontinuous 10 to 15% SDS-PAGE. Numbers on the right indicate molecular masses in kDa. The experiment was repeated three times with similar patterns. (C) Western blots of T. cruzi lysates (left) and TGFL (sample analogous to leftmost lane in panel A) (right) probed with PDNF/ trans -sialidase-specific monoclonal antibody TCN-2. (D) ELISA. TGFL isolation was similar to that for panel A except that TGFL was eluted from GFRα-1-Fc/protein G-Sepharose beads with 0.5 M NaCl-10 mM Tris, pH 7.5, diluted to 0.15 M NaCl, and concentrated by ultrafiltration. TGFL was repurified and eluted as above, stored in 30% glycerol, and then used for an ELISA (microtiter plates were coated with 95 ng/ml TGFL overnight at 4°C, blocked with 5% goat serum for 1 h at 37°C followed by the indicated concentrations of Fc chimera receptors in 5% BSA-PBST, pH 7.2, for 2 h at 37°C, washed, and developed with alkaline phosphatase-labeled anti-human Fc antibody). Points are averages of triplicate points. The experiment was repeated three times with similar results.
    Figure Legend Snippet: Identification and specificity of TGFL. (A) PcIP. The indicated growth factor receptor-Fc chimeras (GFRα-1, RET, TGF-β-RII, FGF-R1α, TNF-RI, and P75) (each, 0.5 μg) were incubated overnight 4°C with T. cruzi trypomastigote lysate (∼25 to 30 μg), adsorbed to protein G-Sepharose, washed, eluted with 2% SDS, run on a nonreducing SDS-PAGE gel, blotted onto nitrocellulose, probed with chagasic sera followed by HRP-labeled secondary anti-human IgG antibody, and visualized with the ECL system for 1 second and 20 seconds. Numbers on the right represent molecular masses in kDa. The experiment was repeated more than 15 times with nearly identical results. Control lanes, lysate adsorbed to protein G-Sepharose without premixing with receptor ( T. cruzi +) and receptor adsorbed to protein G-Sepharose without premixing with lysate (receptor +); adsorption of lysate-receptor mixtures to protein G-Sepharose is denoted by appropriate + and − signs in the corresponding lanes. (B) Coomassie blue staining of TGFL (1.9 μg, equivalent to 2.3 × 10 8 T. cruzi trypomastigotes, Silvio strain) twice purified on GFRα-1/protein G-Sepharose column after discontinuous 10 to 15% SDS-PAGE. Numbers on the right indicate molecular masses in kDa. The experiment was repeated three times with similar patterns. (C) Western blots of T. cruzi lysates (left) and TGFL (sample analogous to leftmost lane in panel A) (right) probed with PDNF/ trans -sialidase-specific monoclonal antibody TCN-2. (D) ELISA. TGFL isolation was similar to that for panel A except that TGFL was eluted from GFRα-1-Fc/protein G-Sepharose beads with 0.5 M NaCl-10 mM Tris, pH 7.5, diluted to 0.15 M NaCl, and concentrated by ultrafiltration. TGFL was repurified and eluted as above, stored in 30% glycerol, and then used for an ELISA (microtiter plates were coated with 95 ng/ml TGFL overnight at 4°C, blocked with 5% goat serum for 1 h at 37°C followed by the indicated concentrations of Fc chimera receptors in 5% BSA-PBST, pH 7.2, for 2 h at 37°C, washed, and developed with alkaline phosphatase-labeled anti-human Fc antibody). Points are averages of triplicate points. The experiment was repeated three times with similar results.

    Techniques Used: Incubation, SDS Page, Labeling, Adsorption, Staining, Purification, Western Blot, Enzyme-linked Immunosorbent Assay, Isolation

    13) Product Images from "Aminopeptidases do not directly degrade tau protein"

    Article Title: Aminopeptidases do not directly degrade tau protein

    Journal: Molecular Neurodegeneration

    doi: 10.1186/1750-1326-5-48

    Reaction of Tau with various forms of PSA . Same amount of PSA activity was reacted with Tau 2N4R or Tau 0N4R in 10 mM Tris buffer, pH 7.5, 1 mM DTT at 37°C for 16 h. (a). Reaction mixtures were analyzed by SDS-PAGE, only Tau 2N4R is shown. (b). For quantitation, the intensity of Tau remaining was determined using ImageJ software [ 26 ]. For internal control, a 50% Tau alone (last lane) was run and the band intensity shown to be ~50% of the input Tau (first lane). Further, by analyzing the staining intensity of the PSA bands we calculated that the variation in the amount of PSA added in each lane varied less than 4%. For the GST-PSA lane, both GST-PSA and free PSA were present. Their intensities were combined to get total PSA. * = free GST.
    Figure Legend Snippet: Reaction of Tau with various forms of PSA . Same amount of PSA activity was reacted with Tau 2N4R or Tau 0N4R in 10 mM Tris buffer, pH 7.5, 1 mM DTT at 37°C for 16 h. (a). Reaction mixtures were analyzed by SDS-PAGE, only Tau 2N4R is shown. (b). For quantitation, the intensity of Tau remaining was determined using ImageJ software [ 26 ]. For internal control, a 50% Tau alone (last lane) was run and the band intensity shown to be ~50% of the input Tau (first lane). Further, by analyzing the staining intensity of the PSA bands we calculated that the variation in the amount of PSA added in each lane varied less than 4%. For the GST-PSA lane, both GST-PSA and free PSA were present. Their intensities were combined to get total PSA. * = free GST.

    Techniques Used: Activity Assay, SDS Page, Quantitation Assay, Software, Staining

    Human PSA sf preparation cleaves Tau . (a) Purity of hPSA sf (5 μg). (b) Degradation of Tau by hPSA sf . Tau was incubated with PSA at a molar ratio of 15:1 (Tau:PSA) in 10 mM Tris buffer, pH 7.5, 1 mM DTT at 37°C for 16 h. Reactions containing 1 μg Tau were analyzed by SDS-PAGE on a 12.5% gel. Labeled products and intact Tau were further analyzed by mass spectrometry and the result is shown in Table 1. (c) Reaction products immunoblotted with N-terminal Tau antibody 5A6 (red) or C-terminal Tau antibody A00024 (green). Twenty percent of the reaction mix was loaded in each lane. Intact Tau shows a yellow color due to the red/green overlap. N1 also reacts to A00024, but was overshadowed by higher intensity signal of 5A6. Molecular weight markers in kDa are shown at the left.
    Figure Legend Snippet: Human PSA sf preparation cleaves Tau . (a) Purity of hPSA sf (5 μg). (b) Degradation of Tau by hPSA sf . Tau was incubated with PSA at a molar ratio of 15:1 (Tau:PSA) in 10 mM Tris buffer, pH 7.5, 1 mM DTT at 37°C for 16 h. Reactions containing 1 μg Tau were analyzed by SDS-PAGE on a 12.5% gel. Labeled products and intact Tau were further analyzed by mass spectrometry and the result is shown in Table 1. (c) Reaction products immunoblotted with N-terminal Tau antibody 5A6 (red) or C-terminal Tau antibody A00024 (green). Twenty percent of the reaction mix was loaded in each lane. Intact Tau shows a yellow color due to the red/green overlap. N1 also reacts to A00024, but was overshadowed by higher intensity signal of 5A6. Molecular weight markers in kDa are shown at the left.

    Techniques Used: Incubation, SDS Page, Labeling, Mass Spectrometry, Molecular Weight

    14) Product Images from "The role of irradiance and C-use strategies in tropical macroalgae photosynthetic response to ocean acidification"

    Article Title: The role of irradiance and C-use strategies in tropical macroalgae photosynthetic response to ocean acidification

    Journal: Scientific Reports

    doi: 10.1038/s41598-018-27333-0

    Inhibition of P gmax in three fleshy ( a ) and five calcifying ( b ) tropical macroalgae as inhibitor block various bicarbonate uptake pathways: external carbonic anhydrase (CA ext ) with acetazolamide (AZ), AE protein by pyridoxal (5) phosphate (PLP), proton pump acidification by Tris buffer and ATPase H + pumps by sodium orthovanadate.
    Figure Legend Snippet: Inhibition of P gmax in three fleshy ( a ) and five calcifying ( b ) tropical macroalgae as inhibitor block various bicarbonate uptake pathways: external carbonic anhydrase (CA ext ) with acetazolamide (AZ), AE protein by pyridoxal (5) phosphate (PLP), proton pump acidification by Tris buffer and ATPase H + pumps by sodium orthovanadate.

    Techniques Used: Inhibition, Blocking Assay, Plasmid Purification

    15) Product Images from "The HSV-1 ICP27 RGG box specifically binds flexible, GC-rich sequences but not G-quartet structures"

    Article Title: The HSV-1 ICP27 RGG box specifically binds flexible, GC-rich sequences but not G-quartet structures

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkp793

    1D 1 H spectra of G-quartet DNA and selected gC 30-mer DNA oligonucleotides. A portion of the 1D 1 H spectra (9–14 p.p.m.) collected on 0.5 mM G4 DNA [TTGGGGTT] 4, gC 191–220, gC 91–120, gC 31–60, gC 11–40 and gC 1–30 at 25°C in 50 mM Tris pH 8, 150 mM KCl and 10% D 2 O. Spectra were scaled using a resolved H1’ ribose signal. Asterisks (*) indicate prominent peaks detected in gC 191–220 and gC 31–60 sequences, two sequences that were not shifted by the ICP27 in EMSA experiments ( Figure 2 ).
    Figure Legend Snippet: 1D 1 H spectra of G-quartet DNA and selected gC 30-mer DNA oligonucleotides. A portion of the 1D 1 H spectra (9–14 p.p.m.) collected on 0.5 mM G4 DNA [TTGGGGTT] 4, gC 191–220, gC 91–120, gC 31–60, gC 11–40 and gC 1–30 at 25°C in 50 mM Tris pH 8, 150 mM KCl and 10% D 2 O. Spectra were scaled using a resolved H1’ ribose signal. Asterisks (*) indicate prominent peaks detected in gC 191–220 and gC 31–60 sequences, two sequences that were not shifted by the ICP27 in EMSA experiments ( Figure 2 ).

    Techniques Used:

    1D 1 H spectra of G-quartet DNA and selected SELEX DNA sequences. A portion of the 1D 1 H spectra (9–14 p.p.m.) collected on 0.5 mM G4 DNA [dTTGGGGTT] 4, SELEX 13, SELEX 4 and SELEX 2 at 25°C in 50 mM Tris pH 8, 150 mM KCl and 10% D 2 O. Spectra were scaled using a resolved H1’ ribose signal. The asterisk (*) indicates broad peaks specifically seen in SELEX 2, a sequence that was not shifted by the ICP27 in EMSA experiments ( Figure 6 ).
    Figure Legend Snippet: 1D 1 H spectra of G-quartet DNA and selected SELEX DNA sequences. A portion of the 1D 1 H spectra (9–14 p.p.m.) collected on 0.5 mM G4 DNA [dTTGGGGTT] 4, SELEX 13, SELEX 4 and SELEX 2 at 25°C in 50 mM Tris pH 8, 150 mM KCl and 10% D 2 O. Spectra were scaled using a resolved H1’ ribose signal. The asterisk (*) indicates broad peaks specifically seen in SELEX 2, a sequence that was not shifted by the ICP27 in EMSA experiments ( Figure 6 ).

    Techniques Used: Sequencing

    EMSA of selected SELEX DNA oligonucleotides. A 20 fmol of radiolabeled SELEX 20-mer DNA oligonucleotides (see Table 2 ) or the gC 11–40 DNA oligonucleotide were either incubated no protein or with 2.5–62.5 µM of the ICP27 N-terminal peptide. A 62.5 µM of the ICP27 N-terminal peptide was used in the (+) gC 11–40 lanes. Samples were electrophoresed on a prerun acrylamide gel with Tris Acetate Buffer and dried gels were exposed to film. Arrows indicate the migration of free probe and the shift due to ICP27 N-terminal peptide binding.
    Figure Legend Snippet: EMSA of selected SELEX DNA oligonucleotides. A 20 fmol of radiolabeled SELEX 20-mer DNA oligonucleotides (see Table 2 ) or the gC 11–40 DNA oligonucleotide were either incubated no protein or with 2.5–62.5 µM of the ICP27 N-terminal peptide. A 62.5 µM of the ICP27 N-terminal peptide was used in the (+) gC 11–40 lanes. Samples were electrophoresed on a prerun acrylamide gel with Tris Acetate Buffer and dried gels were exposed to film. Arrows indicate the migration of free probe and the shift due to ICP27 N-terminal peptide binding.

    Techniques Used: Incubation, Acrylamide Gel Assay, Migration, Binding Assay

    16) Product Images from "In vitro Conversion of Vinyl to Formyl Groups in Naturally Occurring Chlorophylls"

    Article Title: In vitro Conversion of Vinyl to Formyl Groups in Naturally Occurring Chlorophylls

    Journal: Scientific Reports

    doi: 10.1038/srep06069

    pH dependence of Chl d chemical synthesis from Chl a . Chromatograms (spectrum maximum plots) of detergent solubilised Chl a reacted with β-mercaptoethanol and heme in a 50 mM Tris buffer at the indicated pH. Unreacted Chl a , Chl d and 3 1 -sulfoxide of β-mercaptoethanol derivatives (a_I and a_II) are marked. Proportions of the major products and remaining Chl a and Chl a ’ (the C13 2 -Chl a epimer) are indicated in the pie charts for each pH tested.
    Figure Legend Snippet: pH dependence of Chl d chemical synthesis from Chl a . Chromatograms (spectrum maximum plots) of detergent solubilised Chl a reacted with β-mercaptoethanol and heme in a 50 mM Tris buffer at the indicated pH. Unreacted Chl a , Chl d and 3 1 -sulfoxide of β-mercaptoethanol derivatives (a_I and a_II) are marked. Proportions of the major products and remaining Chl a and Chl a ’ (the C13 2 -Chl a epimer) are indicated in the pie charts for each pH tested.

    Techniques Used:

    17) Product Images from "Mechanism of Reconstitution/Activation of the Soluble PQQ-Dependent Glucose Dehydrogenase from Acinetobactercalcoaceticus: A Comprehensive Study"

    Article Title: Mechanism of Reconstitution/Activation of the Soluble PQQ-Dependent Glucose Dehydrogenase from Acinetobactercalcoaceticus: A Comprehensive Study

    Journal: ACS Omega

    doi: 10.1021/acsomega.9b04034

    Transient traces of the quenching of tryptophan fluorescence residues of apo-sGDH during enzyme reconstitution with PQQ and calcium. (A) Stopped-flow kinetics obtained after mixing 2.5 μM subunits of apo-sGDH with 3 mM CaCl 2 and different concentrations of PQQ (from left to right): 40, 25, 15, 10, 5, and 2.5 μM. All concentrations given are after mixing. The reactions were carried out at 10 °C in a 50 mM Tris buffer (pH 7.5), and the fluorescence was monitored at 340 nm with an excitation at 297 nm. (B) Stopped-flow kinetics obtained after mixing 2.5 μM subunits of apo-sGDH with 5 μM PQQ and different concentrations of CaCl 2 (from left to right): 9 μM, 100 μM, 3 mM, 30 mM, 60 mM, and 150 mM. All concentrations given are after mixing, and other experimental conditions are the same as in (A). The black dotted curves in (A) and (B) are the best fits of a monoexponential function to the experimental data.
    Figure Legend Snippet: Transient traces of the quenching of tryptophan fluorescence residues of apo-sGDH during enzyme reconstitution with PQQ and calcium. (A) Stopped-flow kinetics obtained after mixing 2.5 μM subunits of apo-sGDH with 3 mM CaCl 2 and different concentrations of PQQ (from left to right): 40, 25, 15, 10, 5, and 2.5 μM. All concentrations given are after mixing. The reactions were carried out at 10 °C in a 50 mM Tris buffer (pH 7.5), and the fluorescence was monitored at 340 nm with an excitation at 297 nm. (B) Stopped-flow kinetics obtained after mixing 2.5 μM subunits of apo-sGDH with 5 μM PQQ and different concentrations of CaCl 2 (from left to right): 9 μM, 100 μM, 3 mM, 30 mM, 60 mM, and 150 mM. All concentrations given are after mixing, and other experimental conditions are the same as in (A). The black dotted curves in (A) and (B) are the best fits of a monoexponential function to the experimental data.

    Techniques Used: Fluorescence

    Stopped-flow transient traces of the catalytic reduction of PQQ by glucose during the reconstitution/activation of apo-sGDH into holo-sGDH. (A) Kinetic traces (average of four consecutive experiments) obtained after mixing 2.5 μM subunits of apo-sGDH with 100 μM glucose, 3 mM CaCl 2 , and different concentrations of PQQ (from left to right): 40, 25, 15, 10, 5, 2.5, 2, 1.5, 1, and 0.5 μM (the code color for each [PQQ] is reported on the graph). All concentrations given are after mixing. The reactions were monitored at 338 nm and 10 °C in a 50 mM Tris buffer (pH 7.5). (B) Titration plot of apo-GDH by PQQ obtained from the plot of total absorbance change in (A) as a function of the [PQQ]/[apo-sGDH] ratio. (C) Kinetic traces (average of four consecutive experiments) obtained after mixing 2.5 μM subunits of apo-sGDH with 5 μM PQQ (2-fold excess), 100 μM glucose, and different concentrations of CaCl 2 : 9 μM, 100 μM, 3 mM, 30 mM, 60 mM, and 150 mM (the code color for each [CaCl 2 ] is reported on the graph). All concentrations given are after mixing, and other experimental conditions are the same as in (A). The black dotted curves in (A) and (C) are the best fits of a biexponential law to the experimental data.
    Figure Legend Snippet: Stopped-flow transient traces of the catalytic reduction of PQQ by glucose during the reconstitution/activation of apo-sGDH into holo-sGDH. (A) Kinetic traces (average of four consecutive experiments) obtained after mixing 2.5 μM subunits of apo-sGDH with 100 μM glucose, 3 mM CaCl 2 , and different concentrations of PQQ (from left to right): 40, 25, 15, 10, 5, 2.5, 2, 1.5, 1, and 0.5 μM (the code color for each [PQQ] is reported on the graph). All concentrations given are after mixing. The reactions were monitored at 338 nm and 10 °C in a 50 mM Tris buffer (pH 7.5). (B) Titration plot of apo-GDH by PQQ obtained from the plot of total absorbance change in (A) as a function of the [PQQ]/[apo-sGDH] ratio. (C) Kinetic traces (average of four consecutive experiments) obtained after mixing 2.5 μM subunits of apo-sGDH with 5 μM PQQ (2-fold excess), 100 μM glucose, and different concentrations of CaCl 2 : 9 μM, 100 μM, 3 mM, 30 mM, 60 mM, and 150 mM (the code color for each [CaCl 2 ] is reported on the graph). All concentrations given are after mixing, and other experimental conditions are the same as in (A). The black dotted curves in (A) and (C) are the best fits of a biexponential law to the experimental data.

    Techniques Used: Activation Assay, Titration

    18) Product Images from "Long-Lived States of Magnetically Equivalent Spins Populated by Dissolution-DNP and Revealed by Enzymatic Reactions **"

    Article Title: Long-Lived States of Magnetically Equivalent Spins Populated by Dissolution-DNP and Revealed by Enzymatic Reactions **

    Journal: Chemistry (Weinheim an Der Bergstrasse, Germany)

    doi: 10.1002/chem.201404967

    Enzymatic conversion of fumarate into malate by fumarase at 300 K monitored by integration of the conventional 1 H NMR signals of the two species. The nuclear polarizations are in thermal Boltzmann equilibrium, without resorting to DNP. A solution of fumarase (4 μL, 5.8 mg mL −1 , i.e., 10 units) was injected into a fumarate solution (500 μL, 50 m M ) at pH 8 in a buffer of 25 m M TRIS and 200 m M NaCl.
    Figure Legend Snippet: Enzymatic conversion of fumarate into malate by fumarase at 300 K monitored by integration of the conventional 1 H NMR signals of the two species. The nuclear polarizations are in thermal Boltzmann equilibrium, without resorting to DNP. A solution of fumarase (4 μL, 5.8 mg mL −1 , i.e., 10 units) was injected into a fumarate solution (500 μL, 50 m M ) at pH 8 in a buffer of 25 m M TRIS and 200 m M NaCl.

    Techniques Used: Nuclear Magnetic Resonance, Injection

    Related Articles

    Polymerase Chain Reaction:

    Article Title: Genome-Wide and Gene-Specific Epigenomic Platforms for Hepatocellular Carcinoma Biomarker Development Trials
    Article Snippet: .. Fluorogenic PCR reactions were carried out in a reaction volume of 20 μ L consisting of 600 nmol/L of each primer, 200 μ mol/L probe, 0.75 units platinum Taq polymerase (Invitrogen), 200 μ mol/L of each dATP, dCTP, dGTP, and dTTP, 200 nmol/L ROX dye reference (Invitrogen), 16.6 mmol/L ammonium sulfate, 67 mmol/L Trizma (Sigma, St. Louis, MO), 6.7 mmol/L magnesium chloride, 10 mmol/L mercaptoethanol, and 0.1% DMSO. .. Duplicates of three microliters (3 μ L) of bisulfite-modified DNA solution were used in each real-time methylation-specific PCR (qMSP) amplification reaction.

    Plasmid Purification:

    Article Title: The role of irradiance and C-use strategies in tropical macroalgae photosynthetic response to ocean acidification
    Article Snippet: .. Inhibitors included acetazolamide (AZ, Sigma Aldrich) that blocks the dehydration of HCO3 − into CO2 via external carbonate anhydrase (CAext ) , pyridoxal (5) phosphate (PLP, Fisher Scientific) that inhibits active uptake of HCO3 − , Tris buffer (Trizma R, Sigma Aldrich) that interferes with proton pump acidification of the thalli boundary layer and sodium orthovanadate (vanadate, Sigma Aldrich) that obstructs plasmalemma ATPase H+ pumps . .. Solutions of AZ (200 μM), PLP (480 μM) and Tris (50 mM) were dissolved in filtered seawater (0.45 μm) followed by pH adjustment to 8.1 (1 M HCl).

    Incubation:

    Article Title: Eggplant Resistance to the Ralstonia solanacearum Species Complex Involves Both Broad-Spectrum and Strain-Specific Quantitative Trait Loci
    Article Snippet: .. Actively growing cultures were harvested after a 24 h incubation period, by flooding plates with 5 mL of Tris buffer (Trizma 0.01 M pH 7.2: Sigma, St Louis, MO, United States). .. The bacterial suspension was then titrated by spectrophotometry to adjust the inoculum concentration at approximately 1 × 108 CFU per mL (OD600nm = 0.1).

    SYBR Green Assay:

    Article Title: Effect of Sodium Valproate on the Toxicity of Cyclophosphamide in the Testes of Mice: Influence of Pre- and Post-Treatment Schedule
    Article Snippet: .. Chemicals Sodium valproate (CAS no. 1069-66-5), cyclophosphamide (CAS no. 6055-19-2), bovine serum albumin (CAS no. 9048-46-8), hematoxylin and eosin (H and E), Trizma (CAS no. 77-86-1), dithiothreitol (CAS no. 3483-12-3), Proteinase-K (CAS no. 39450-01-6) and SYBR Green I (CAS no. 163795-75-3) were purchased from Sigma-Aldrich Chemicals, Saint Louis, MO, USA. .. Dimethyl sulphoxide (DMSO), normal melting point agarose (NMPA), low melting point agarose (LMPA), Triton X-100, ethylenediamine-tetraacetic acid (EDTA) and Hank's balanced salt solution (HBSS) were obtained from HiMedia Laboratories Ltd., Mumbai.

    Concentration Assay:

    Article Title: Real-time cardiac metabolism assessed with hyperpolarized [1-13C]acetate in a large-animal model
    Article Snippet: .. C-1 relaxivity was also determined at 9.4 T ( VnmrS , Agilent Technologies, Santa Clara, CA, USA) for 20% enriched Na[1-13 C]acetate samples with increased trityl radical concentration (0.44 mM, 2.5 mM and 5 mM respectively, 13 C-acetate final concentration 90 mM) in a 40 mM Trizma® (Sigma-Aldrich), 0.27 mM EDTA (Sigma-Aldrich) buffer. .. T1 was determined from a mono-exponential fit using an Inversion Recovery (IR) sequence; r1 was thus estimated at 9.4 T and 25 °C from the linear fit of the relaxation rate (R1 ) as a function of the OX063 concentration.

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  • 99
    Millipore tbe buffer
    Analysis of heterotrimerization. Purified recombinant proteins were separated by 4% native PAGE in 1× <t>TBE</t> buffer at 4°C. Lane 1: His At PCNA1; lane 2: At PCNA1; lane 3: His At <t>PCNA2;</t> lane 4: At PCNA2; lane 5: His At PCNA1 and At PCNA2 mixed after
    Tbe Buffer, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 18 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore tris buffered saline
    ( A ) Fibrinolytic effects of rSKs on human plasma clots. Human plasma (45 μl) was mixed with trace amounts of 125 ). After a wash with <t>Tris-buffered</t> saline (pH 7.4) containing the thrombin inhibitor <t>PPACK</t> (10 μM), the clots were suspended in 2 ml of plasma containing 10 μM PPACK. Various amounts of rSK and rSKΔ59 that had undergone active-site titration (0–50 nM) were added to the plasma, and, after 6 h, the amount of fibrinolysis was determined by measuring the release of soluble 125 ). The means ± SD are shown (but in some cases the SD are too small to be visible). ( B ) Fibrinogen degradation by rSK activator complexes. At the end of the fibrinolysis experiments presented in A ). The means ± SD are shown (but in some cases the SD are too small to be visible).
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    95
    Millipore tris glycine sds page
    Iron chelators inhibit phosphorylation of RNAPII ( a ) Chelators do not have an effect on expression of CDK9 . 293 cells were treated for 24 h with 10 μM 311 or 100 μM of ICL670. The cells were lysed were resolved on 10% <t>SDS-PAGE,</t> and immunoblotted anti-CDK9 antibodies. Lane 1 : control untreated cells; lanes 2, 3 and 4 : cells treated with DFO, 311 or ICL670. ( b ) Chelators inhibit association of CDK9 with cyclin T1 . The cell lysates prepared as in ( a ) were subjected to immunoprecipitation with anti-cyclin T1 antibody (lanes 3–6). The immunoprecipitated material was resolved on 10% SDS-PAGE, and immunoblotted anti-CDK9 antibodies. Lane 1 : input control untreated cells; lane 2: precipitation of untreated cells with non-specific preimmune IgG, lanes 3–6: immunoprecipitation with anti-cyclin T1 antibodies of lysates from untreated, DFO, 311 and ICL670 treated cells. The precipitated samples were resolved on 10% SDS-PAGE and immunoblotted with antibodies against CDK9. ( c ) Chelators inhibit phosphorylation of RNAPII . The cell lysates prepared as in ( a ) were resolved on 7.5% SDS-PAGE, and immunoblotted with the RNAPII CTD phospho-Serine 2 specific antibodies. Lane 1 : control untreated cells; lanes 2,3 and 4 : cells treated with DFO, 311 or ICL670.
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    Analysis of heterotrimerization. Purified recombinant proteins were separated by 4% native PAGE in 1× TBE buffer at 4°C. Lane 1: His At PCNA1; lane 2: At PCNA1; lane 3: His At PCNA2; lane 4: At PCNA2; lane 5: His At PCNA1 and At PCNA2 mixed after

    Journal: Protein Science : A Publication of the Protein Society

    Article Title: Crystal structures of the Arabidopsis thaliana proliferating cell nuclear antigen 1 and 2 proteins complexed with the human p21 C-terminal segment

    doi: 10.1002/pro.117

    Figure Lengend Snippet: Analysis of heterotrimerization. Purified recombinant proteins were separated by 4% native PAGE in 1× TBE buffer at 4°C. Lane 1: His At PCNA1; lane 2: At PCNA1; lane 3: His At PCNA2; lane 4: At PCNA2; lane 5: His At PCNA1 and At PCNA2 mixed after

    Article Snippet: His At PCNA1, At PCNA1, His At PCNA2, At PCNA2, and the heterotrimer were separated by 4% native PAGE in 1× TBE buffer and electrotransferred onto a PVDF membrane (0.2 μm) (Millipore), as described previously.

    Techniques: Purification, Recombinant, Clear Native PAGE

    ( A ) Fibrinolytic effects of rSKs on human plasma clots. Human plasma (45 μl) was mixed with trace amounts of 125 ). After a wash with Tris-buffered saline (pH 7.4) containing the thrombin inhibitor PPACK (10 μM), the clots were suspended in 2 ml of plasma containing 10 μM PPACK. Various amounts of rSK and rSKΔ59 that had undergone active-site titration (0–50 nM) were added to the plasma, and, after 6 h, the amount of fibrinolysis was determined by measuring the release of soluble 125 ). The means ± SD are shown (but in some cases the SD are too small to be visible). ( B ) Fibrinogen degradation by rSK activator complexes. At the end of the fibrinolysis experiments presented in A ). The means ± SD are shown (but in some cases the SD are too small to be visible).

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    Article Title: A catalytic switch and the conversion of streptokinase to a fibrin-targeted plasminogen activator

    doi:

    Figure Lengend Snippet: ( A ) Fibrinolytic effects of rSKs on human plasma clots. Human plasma (45 μl) was mixed with trace amounts of 125 ). After a wash with Tris-buffered saline (pH 7.4) containing the thrombin inhibitor PPACK (10 μM), the clots were suspended in 2 ml of plasma containing 10 μM PPACK. Various amounts of rSK and rSKΔ59 that had undergone active-site titration (0–50 nM) were added to the plasma, and, after 6 h, the amount of fibrinolysis was determined by measuring the release of soluble 125 ). The means ± SD are shown (but in some cases the SD are too small to be visible). ( B ) Fibrinogen degradation by rSK activator complexes. At the end of the fibrinolysis experiments presented in A ). The means ± SD are shown (but in some cases the SD are too small to be visible).

    Article Snippet: After a wash with Tris-buffered saline (pH 7.4) containing the thrombin inhibitor PPACK (10 μM, d -Phe-Pro-Arg chloromethylketone, Calbiochem), the clots were suspended in 2 ml of plasma containing 10 μM PPACK.

    Techniques: Titration

    Iron chelators inhibit phosphorylation of RNAPII ( a ) Chelators do not have an effect on expression of CDK9 . 293 cells were treated for 24 h with 10 μM 311 or 100 μM of ICL670. The cells were lysed were resolved on 10% SDS-PAGE, and immunoblotted anti-CDK9 antibodies. Lane 1 : control untreated cells; lanes 2, 3 and 4 : cells treated with DFO, 311 or ICL670. ( b ) Chelators inhibit association of CDK9 with cyclin T1 . The cell lysates prepared as in ( a ) were subjected to immunoprecipitation with anti-cyclin T1 antibody (lanes 3–6). The immunoprecipitated material was resolved on 10% SDS-PAGE, and immunoblotted anti-CDK9 antibodies. Lane 1 : input control untreated cells; lane 2: precipitation of untreated cells with non-specific preimmune IgG, lanes 3–6: immunoprecipitation with anti-cyclin T1 antibodies of lysates from untreated, DFO, 311 and ICL670 treated cells. The precipitated samples were resolved on 10% SDS-PAGE and immunoblotted with antibodies against CDK9. ( c ) Chelators inhibit phosphorylation of RNAPII . The cell lysates prepared as in ( a ) were resolved on 7.5% SDS-PAGE, and immunoblotted with the RNAPII CTD phospho-Serine 2 specific antibodies. Lane 1 : control untreated cells; lanes 2,3 and 4 : cells treated with DFO, 311 or ICL670.

    Journal: Virology

    Article Title: Iron Chelators ICL670 and 311 Inhibit HIV-1 Transcription

    doi: 10.1016/j.virol.2007.06.011

    Figure Lengend Snippet: Iron chelators inhibit phosphorylation of RNAPII ( a ) Chelators do not have an effect on expression of CDK9 . 293 cells were treated for 24 h with 10 μM 311 or 100 μM of ICL670. The cells were lysed were resolved on 10% SDS-PAGE, and immunoblotted anti-CDK9 antibodies. Lane 1 : control untreated cells; lanes 2, 3 and 4 : cells treated with DFO, 311 or ICL670. ( b ) Chelators inhibit association of CDK9 with cyclin T1 . The cell lysates prepared as in ( a ) were subjected to immunoprecipitation with anti-cyclin T1 antibody (lanes 3–6). The immunoprecipitated material was resolved on 10% SDS-PAGE, and immunoblotted anti-CDK9 antibodies. Lane 1 : input control untreated cells; lane 2: precipitation of untreated cells with non-specific preimmune IgG, lanes 3–6: immunoprecipitation with anti-cyclin T1 antibodies of lysates from untreated, DFO, 311 and ICL670 treated cells. The precipitated samples were resolved on 10% SDS-PAGE and immunoblotted with antibodies against CDK9. ( c ) Chelators inhibit phosphorylation of RNAPII . The cell lysates prepared as in ( a ) were resolved on 7.5% SDS-PAGE, and immunoblotted with the RNAPII CTD phospho-Serine 2 specific antibodies. Lane 1 : control untreated cells; lanes 2,3 and 4 : cells treated with DFO, 311 or ICL670.

    Article Snippet: In parallel, the immunoprecipitated CDK2 was resolved on 10% Tris-Glycine SDS-PAGE, transferred to polyvinylidene fluoride (PVDF) membranes (Millipore, Allen, TX) and immunoblotted with anti-CDK2 antibodies.

    Techniques: Expressing, SDS Page, Immunoprecipitation

    Iron chelators reduce HIV-1 Tat phosphorylated in cultured cells HeLa cells were infected with recombinant adenovirus expressing Flag-tagged Tat as described in Methods (lanes 2–5). Lane 1, control uninfected cells. HeLa cells were transfected with siRNAs targeting CDK2 (lane 3) or treated with 10 μM 311 or 100 μM ICL670. At 48 hours post infection cells were labeled with ( 32 P)-orthophosphate for 2 hours with the addition of 1 μM okadaic acid. Whole cell extracts were prepared and Tat was immunoprecipitated with anti-Flag monoclonal antibodies, resolved on 15% Tris-Tricine SDS-PAGE and detected by Phosphor Imager. Phosphor Imager quantification is shown in the lower panel.

    Journal: Virology

    Article Title: Iron Chelators ICL670 and 311 Inhibit HIV-1 Transcription

    doi: 10.1016/j.virol.2007.06.011

    Figure Lengend Snippet: Iron chelators reduce HIV-1 Tat phosphorylated in cultured cells HeLa cells were infected with recombinant adenovirus expressing Flag-tagged Tat as described in Methods (lanes 2–5). Lane 1, control uninfected cells. HeLa cells were transfected with siRNAs targeting CDK2 (lane 3) or treated with 10 μM 311 or 100 μM ICL670. At 48 hours post infection cells were labeled with ( 32 P)-orthophosphate for 2 hours with the addition of 1 μM okadaic acid. Whole cell extracts were prepared and Tat was immunoprecipitated with anti-Flag monoclonal antibodies, resolved on 15% Tris-Tricine SDS-PAGE and detected by Phosphor Imager. Phosphor Imager quantification is shown in the lower panel.

    Article Snippet: In parallel, the immunoprecipitated CDK2 was resolved on 10% Tris-Glycine SDS-PAGE, transferred to polyvinylidene fluoride (PVDF) membranes (Millipore, Allen, TX) and immunoblotted with anti-CDK2 antibodies.

    Techniques: Cell Culture, Infection, Recombinant, Expressing, Transfection, Labeling, Immunoprecipitation, SDS Page

    Iron chelators reduce cellular activity of CDK2 (a) CDK2 expression determined by Western blotting . 293 cells were grown in 100 mm plates and treated for the indicated time with 100 μM DFO, 10 μM 311 or 100 μM ICL670. The cells were lysed and CDK2 was immunoprecipitated as described in Methods. Lane 1, input control. Lane, preimmune IgG was used for the immunoprecipitation. The precipitated CDK2 was resolved on 10% SDS-PAGE and immunoblotted with antibodies against CDK2. ( b ) CDK2 was precipitated as in (a) and the immunoprecipitated material was subsequently incubated with histone H1 in the presence of γ-( 32 P) ATP. Kinase reactions were resolved on 10% SDS-PAGE and analyzed on Phosphor Imager. Lane 1 : non-specific preimmune IgG. Lane 10, histone H1 phosphorylation with recombinant CDK2/Cyclin E.

    Journal: Virology

    Article Title: Iron Chelators ICL670 and 311 Inhibit HIV-1 Transcription

    doi: 10.1016/j.virol.2007.06.011

    Figure Lengend Snippet: Iron chelators reduce cellular activity of CDK2 (a) CDK2 expression determined by Western blotting . 293 cells were grown in 100 mm plates and treated for the indicated time with 100 μM DFO, 10 μM 311 or 100 μM ICL670. The cells were lysed and CDK2 was immunoprecipitated as described in Methods. Lane 1, input control. Lane, preimmune IgG was used for the immunoprecipitation. The precipitated CDK2 was resolved on 10% SDS-PAGE and immunoblotted with antibodies against CDK2. ( b ) CDK2 was precipitated as in (a) and the immunoprecipitated material was subsequently incubated with histone H1 in the presence of γ-( 32 P) ATP. Kinase reactions were resolved on 10% SDS-PAGE and analyzed on Phosphor Imager. Lane 1 : non-specific preimmune IgG. Lane 10, histone H1 phosphorylation with recombinant CDK2/Cyclin E.

    Article Snippet: In parallel, the immunoprecipitated CDK2 was resolved on 10% Tris-Glycine SDS-PAGE, transferred to polyvinylidene fluoride (PVDF) membranes (Millipore, Allen, TX) and immunoblotted with anti-CDK2 antibodies.

    Techniques: Activity Assay, Expressing, Western Blot, Immunoprecipitation, SDS Page, Incubation, Recombinant

    Resolution of OMPs from E. aerogenes type strain ATCC 13048, E. aerogenes 810, and E. aerogenes 810-REV on 4 to 16% (A) and 4 to 12% (B) Tris-glycine-SDS gels and 4 to 12% NuPAGE gels (C). Strain 810 was grown in the presence and absence of imipenem. The closed arrowheads indicate the 42-kDa porin (presumed OmpC analog), and the open arrowheads indicate the 39-kDa porin (presumed OmpF analog).

    Journal: Antimicrobial Agents and Chemotherapy

    Article Title: Carbapenem Resistance in a Clinical Isolate of Enterobacter aerogenes Is Associated with Decreased Expression of OmpF and OmpC Porin Analogs

    doi: 10.1128/AAC.46.12.3817-3822.2002

    Figure Lengend Snippet: Resolution of OMPs from E. aerogenes type strain ATCC 13048, E. aerogenes 810, and E. aerogenes 810-REV on 4 to 16% (A) and 4 to 12% (B) Tris-glycine-SDS gels and 4 to 12% NuPAGE gels (C). Strain 810 was grown in the presence and absence of imipenem. The closed arrowheads indicate the 42-kDa porin (presumed OmpC analog), and the open arrowheads indicate the 39-kDa porin (presumed OmpF analog).

    Article Snippet: Western blotting of porins was performed as follows: proteins from 4 to 12% Tris-glycine-SDS gels or 10% NuPAGE gels were transferred to Immobilon-P filters (Millipore) ( , ).

    Techniques: