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Boehringer Mannheim phenylmethylsulfonyl fluoride
Phenylmethylsulfonyl Fluoride, supplied by Boehringer Mannheim, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Nucleic Acid Electrophoresis:

Article Title: Potentiation of Excitotoxicity in Transgenic Mice Overexpressing Neuronal Cyclooxygenase-2
Article Snippet: .. Brain samples were homogenized in an ice-cold buffer of Tris, pH 7.2, containing 1% NP-40, 0.5% sodium deoxycholate, 0.1% lauryl sulfate, 100 μg/ml phenylmethylsulfonyl fluorid, 3 μg/ml aprotinin, 3 μg/ml pepstatin A, 15 μg/ml propionyl-Leupeptin, and 600 μg/ml Perfabloc (Boehringer Mannheim, Indianapolis, IN), boiled, centrifuged, and electrophoresed on 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) as previously described. .. Proteins were transferred to nylon Transblot membrane (Biorad, Hercules, CA) and immunoreacted with a polyclonal goat anti-hCOX-2 antibody (1:500 dilution, 3 hours, room temperature) in Superblock blocking solution (Pierce, Rockford, IL).

SDS Page:

Article Title: Potentiation of Excitotoxicity in Transgenic Mice Overexpressing Neuronal Cyclooxygenase-2
Article Snippet: .. Brain samples were homogenized in an ice-cold buffer of Tris, pH 7.2, containing 1% NP-40, 0.5% sodium deoxycholate, 0.1% lauryl sulfate, 100 μg/ml phenylmethylsulfonyl fluorid, 3 μg/ml aprotinin, 3 μg/ml pepstatin A, 15 μg/ml propionyl-Leupeptin, and 600 μg/ml Perfabloc (Boehringer Mannheim, Indianapolis, IN), boiled, centrifuged, and electrophoresed on 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) as previously described. .. Proteins were transferred to nylon Transblot membrane (Biorad, Hercules, CA) and immunoreacted with a polyclonal goat anti-hCOX-2 antibody (1:500 dilution, 3 hours, room temperature) in Superblock blocking solution (Pierce, Rockford, IL).

Protease Inhibitor:

Article Title: Expression of Nuclear Factor Erythroid 2 Protein in Malignant Cutaneous Tumors
Article Snippet: Western blot analysis The human MM cell line G361 served as a positive control for Nrf2 expression. .. The tissue samples were homogenized in WCE buffer (25 mM HEPES [pH 7.7], 0.3 M NaCl, 1.5 mM MgCl2 , 0.2 mM ethylene diamine tetraacetic acid [EDTA], 0.1% Triton X-100, 0.5 mM dithiothreitol [DTT], 20 mM glycerophosphate, 0.1 mM Na3 VO4 , 2 g per mL leupeptin, 2 g per mL aprotinin, 1 mM phenylmethylsulfonyl fluoride [PMSF], and a protease inhibitor cocktail tablet [Boehringer Mannheim, Mannheim, Germany]). ..

Article Title: Identification of a Translation Initiation Factor 3 (eIF3) Core Complex, Conserved in Yeast and Mammals, That Interacts with eIF5
Article Snippet: Cells were grown in 10 liters of YPD medium to an optical density at 600 nm (OD600 ) of 7.5 to 8.0, harvested by centrifugation at 7,000 × g for 15 min, and washed with ice-cold water. .. About 80 g of cells was resuspended in 160 ml of buffer A (20 mM Tris-HCl [pH 7.5], 100 mM KCl, 5 mM MgCl2 , 0.1 mM EDTA, 7 mM β-mercaptoethanol, 1 mM phenylmethylsulfonyl fluoride [PMSF], 1× Complete protease inhibitor cocktail [Boehringer Mannheim]) and homogenized in a bead beater with 2 cell volumes of glass beads. .. The pooled homogenate was clarified by centrifugation at 17,000 × g and then at 25,000 × g for 15 min, and ribosomes were pelleted at 200,000 × g for 2 h. The ribosomal pellet was resuspended in 25 ml of buffer B (buffer A containing 350 mM KCl) and centrifuged at 200,000 × g for 2 h.

Article Title: Cytomegalovirus IE2 Protein Stimulates Interleukin 1? Gene Transcription via Tethering to Spi-1/PU.1
Article Snippet: After 40 h of incubation at 37°C with 5% CO2 , cells were washed with cold PBS once and detached with cold PBS containing 25 mM EDTA. .. The cells were collected by centrifugation and resuspended in 200 μl of cold lysis buffer (20 mM HEPES [pH 7.9], 1 mM MgCl2 , 10 mM KCl, 300 mM NaCl, 0.1% Triton X-100, 20% glycerol, 1 mM dithiothreitol, 1 mM phenylmethylsulfonyl fluoride [PMSF], 10 mM NaF, 1 mM ZnCl2 , 1 mM Na orthovanadate, 1 protease inhibitor cocktail tablet [Boehringer Mannheim]/50 ml). .. Protein concentrations were measured by the Bradford method (Bio-Rad).

Article Title: The Helicase-Like Domain of Plant Potexvirus Replicase Participates in Formation of RNA 5? Cap Structure by Exhibiting RNA 5?-Triphosphatase Activity
Article Snippet: Escherichia coli Novablue cells (Novagen) harboring the desired plasmids were grown in Luria-Bertani broth, and the helicase-like domain was produced by the induction of IPTG (isopropyl-β- d -thiogalactopyranoside). .. Cells were then harvested by centrifugation, suspended in lysis buffer (20 mM Tris [pH 7.4], 100 mM KCl, 0.1% Brij-35, 10% glycerol, 10 mM β-mercaptoethanol, 1 mM phenylmethylsulfonyl fluoride [PMSF], and protease inhibitor cocktail [Boehringer Mannheim]), and broken by ultrasonication. .. The majority of the helicase-like domain was pelleted as inclusion bodies, solubilized in 8 M urea-containing Tris buffer (pH 7.5, 50 mM), and immediately refolded in an excess of lysis buffer.

Article Title: Atomic Hydrogen Surrounded by Water Molecules, H(H2O)m, Modulates Basal and UV-Induced Gene Expressions in Human Skin In Vivo
Article Snippet: Western blot analysis To determine the amount of MMP-1 secreted into the culture media, equal aliquots of conditioned culture media from an equal number of cells were fractionated by 10% SDS PAGE, transferred to a Hybond ECL membrane (Amersham Biosciences, Buckinghamshire, England) and analyzed by Western blotting with a antibody against MMP-1 (Lab Frontier, Seoul, Korea) using enhanced chemiluminescence (Amersham Biosciences). .. To analyze MAPK activation, total cell lysates were prepared in a lysis buffer [25 mM Hepes (pH 7.7), 0.3 M NaCl, 1.5 mM MgCl2 , 0.2 mM ethylenediamine tetraacetic acid (EDTA), 0.1% Triton X-100, 0.5 mM dithiothreitol (DTT), 20 mM β-glycerolphosphate, 0.1 mM Na3 VO4 , 2 µg/ml leupeptin, 2 µg/ml aprotinin, 1 mM phenylmethylsulfonyl fluoride (PMSF), and a protease inhibitor cocktail tablet from Boehringer Mannheim (Indianapolis, IN)]. ..

Article Title: Human Immunodeficiency Virus Type 1 Infection Inhibits Granulocyte-Macrophage Colony-Stimulating Factor-Induced Activation of STAT5A in Human Monocyte-Derived Macrophages
Article Snippet: Immunoblots were incubated in a solution consisting of 2% (wt/vol) SDS, 62.5 mM Tris (pH 6.8), and 100 mM β-mercaptoethanol at 60°C for 30 min before reprobing with anti-STAT5A antibody (Santa Cruz Biotechnology) for 2 h. Membranes were again washed and incubated with goat anti-rabbit antibody conjugated to horseradish peroxidase (DAKO Corporation, Carpinteria, Calif.) before ECL assay using Western blotting detection reagents according to the manufacturer's instructions (Amersham Pharmacia Biotech). .. Nuclear extracts were prepared by resuspending cell pellets in a solution containing 10 mM Tris-HCl (pH 7.8), 2 mM MgCl2 , 5 mM KCl, 10% (vol/vol) glycerol, 1 mM EDTA, 2 mM dithiothreitol (DTT), 2 mM phenylmethylsulfonyl fluoride (PMSF), 4 μg of aprotinin per ml, 1.25 μg of leupeptin per ml, and 1× Complete protease inhibitor (Boehringer Mannheim). ..

Article Title: Nonstructural Protein of Infectious Bursal Disease Virus Inhibits Apoptosis at the Early Stage of Virus Infection
Article Snippet: Briefly, 2 × 106 infected CEF cells were trypsinized and washed once with cold PBS. .. The cells were resuspended in 20 μl high-salt detergent buffer containing 20 mM HEPES, pH 7.9, 350 mM NaCl, 20% (wt/vol) glycerol, 1% (wt/vol) NP-40, 1 mM MgCl2 , 0.5 mM EDTA, 0.1 mM EGTA, 0.5 mM dithiothreitol (DTT), 0.1% phenylmethylsulfonyl fluoride (PMSF), and 1% protease inhibitor cocktail (Boehringer Mannheim, Indianapolis, IN) and incubated on ice for 30 min. ..

Centrifugation:

Article Title: Cytomegalovirus IE2 Protein Stimulates Interleukin 1? Gene Transcription via Tethering to Spi-1/PU.1
Article Snippet: After 40 h of incubation at 37°C with 5% CO2 , cells were washed with cold PBS once and detached with cold PBS containing 25 mM EDTA. .. The cells were collected by centrifugation and resuspended in 200 μl of cold lysis buffer (20 mM HEPES [pH 7.9], 1 mM MgCl2 , 10 mM KCl, 300 mM NaCl, 0.1% Triton X-100, 20% glycerol, 1 mM dithiothreitol, 1 mM phenylmethylsulfonyl fluoride [PMSF], 10 mM NaF, 1 mM ZnCl2 , 1 mM Na orthovanadate, 1 protease inhibitor cocktail tablet [Boehringer Mannheim]/50 ml). .. Protein concentrations were measured by the Bradford method (Bio-Rad).

Article Title: The Helicase-Like Domain of Plant Potexvirus Replicase Participates in Formation of RNA 5? Cap Structure by Exhibiting RNA 5?-Triphosphatase Activity
Article Snippet: Escherichia coli Novablue cells (Novagen) harboring the desired plasmids were grown in Luria-Bertani broth, and the helicase-like domain was produced by the induction of IPTG (isopropyl-β- d -thiogalactopyranoside). .. Cells were then harvested by centrifugation, suspended in lysis buffer (20 mM Tris [pH 7.4], 100 mM KCl, 0.1% Brij-35, 10% glycerol, 10 mM β-mercaptoethanol, 1 mM phenylmethylsulfonyl fluoride [PMSF], and protease inhibitor cocktail [Boehringer Mannheim]), and broken by ultrasonication. .. The majority of the helicase-like domain was pelleted as inclusion bodies, solubilized in 8 M urea-containing Tris buffer (pH 7.5, 50 mM), and immediately refolded in an excess of lysis buffer.

Lysis:

Article Title: Cytomegalovirus IE2 Protein Stimulates Interleukin 1? Gene Transcription via Tethering to Spi-1/PU.1
Article Snippet: After 40 h of incubation at 37°C with 5% CO2 , cells were washed with cold PBS once and detached with cold PBS containing 25 mM EDTA. .. The cells were collected by centrifugation and resuspended in 200 μl of cold lysis buffer (20 mM HEPES [pH 7.9], 1 mM MgCl2 , 10 mM KCl, 300 mM NaCl, 0.1% Triton X-100, 20% glycerol, 1 mM dithiothreitol, 1 mM phenylmethylsulfonyl fluoride [PMSF], 10 mM NaF, 1 mM ZnCl2 , 1 mM Na orthovanadate, 1 protease inhibitor cocktail tablet [Boehringer Mannheim]/50 ml). .. Protein concentrations were measured by the Bradford method (Bio-Rad).

Article Title: The Helicase-Like Domain of Plant Potexvirus Replicase Participates in Formation of RNA 5? Cap Structure by Exhibiting RNA 5?-Triphosphatase Activity
Article Snippet: Escherichia coli Novablue cells (Novagen) harboring the desired plasmids were grown in Luria-Bertani broth, and the helicase-like domain was produced by the induction of IPTG (isopropyl-β- d -thiogalactopyranoside). .. Cells were then harvested by centrifugation, suspended in lysis buffer (20 mM Tris [pH 7.4], 100 mM KCl, 0.1% Brij-35, 10% glycerol, 10 mM β-mercaptoethanol, 1 mM phenylmethylsulfonyl fluoride [PMSF], and protease inhibitor cocktail [Boehringer Mannheim]), and broken by ultrasonication. .. The majority of the helicase-like domain was pelleted as inclusion bodies, solubilized in 8 M urea-containing Tris buffer (pH 7.5, 50 mM), and immediately refolded in an excess of lysis buffer.

Article Title: Atomic Hydrogen Surrounded by Water Molecules, H(H2O)m, Modulates Basal and UV-Induced Gene Expressions in Human Skin In Vivo
Article Snippet: Western blot analysis To determine the amount of MMP-1 secreted into the culture media, equal aliquots of conditioned culture media from an equal number of cells were fractionated by 10% SDS PAGE, transferred to a Hybond ECL membrane (Amersham Biosciences, Buckinghamshire, England) and analyzed by Western blotting with a antibody against MMP-1 (Lab Frontier, Seoul, Korea) using enhanced chemiluminescence (Amersham Biosciences). .. To analyze MAPK activation, total cell lysates were prepared in a lysis buffer [25 mM Hepes (pH 7.7), 0.3 M NaCl, 1.5 mM MgCl2 , 0.2 mM ethylenediamine tetraacetic acid (EDTA), 0.1% Triton X-100, 0.5 mM dithiothreitol (DTT), 20 mM β-glycerolphosphate, 0.1 mM Na3 VO4 , 2 µg/ml leupeptin, 2 µg/ml aprotinin, 1 mM phenylmethylsulfonyl fluoride (PMSF), and a protease inhibitor cocktail tablet from Boehringer Mannheim (Indianapolis, IN)]. ..

Activation Assay:

Article Title: Atomic Hydrogen Surrounded by Water Molecules, H(H2O)m, Modulates Basal and UV-Induced Gene Expressions in Human Skin In Vivo
Article Snippet: Western blot analysis To determine the amount of MMP-1 secreted into the culture media, equal aliquots of conditioned culture media from an equal number of cells were fractionated by 10% SDS PAGE, transferred to a Hybond ECL membrane (Amersham Biosciences, Buckinghamshire, England) and analyzed by Western blotting with a antibody against MMP-1 (Lab Frontier, Seoul, Korea) using enhanced chemiluminescence (Amersham Biosciences). .. To analyze MAPK activation, total cell lysates were prepared in a lysis buffer [25 mM Hepes (pH 7.7), 0.3 M NaCl, 1.5 mM MgCl2 , 0.2 mM ethylenediamine tetraacetic acid (EDTA), 0.1% Triton X-100, 0.5 mM dithiothreitol (DTT), 20 mM β-glycerolphosphate, 0.1 mM Na3 VO4 , 2 µg/ml leupeptin, 2 µg/ml aprotinin, 1 mM phenylmethylsulfonyl fluoride (PMSF), and a protease inhibitor cocktail tablet from Boehringer Mannheim (Indianapolis, IN)]. ..

Incubation:

Article Title: Nonstructural Protein of Infectious Bursal Disease Virus Inhibits Apoptosis at the Early Stage of Virus Infection
Article Snippet: Briefly, 2 × 106 infected CEF cells were trypsinized and washed once with cold PBS. .. The cells were resuspended in 20 μl high-salt detergent buffer containing 20 mM HEPES, pH 7.9, 350 mM NaCl, 20% (wt/vol) glycerol, 1% (wt/vol) NP-40, 1 mM MgCl2 , 0.5 mM EDTA, 0.1 mM EGTA, 0.5 mM dithiothreitol (DTT), 0.1% phenylmethylsulfonyl fluoride (PMSF), and 1% protease inhibitor cocktail (Boehringer Mannheim, Indianapolis, IN) and incubated on ice for 30 min. ..

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  • 86
    Boehringer Mannheim buffer a
    Fig. 3. Ni 2+ affinity purification of intact eIF3 complexes containing eIF1 and eIF5 from heat-treated extract containing His 8 -prt1-1. Translation extracts were prepared from strains LPY201 ( PRT1-His ) and LPY202 ( prt1-1-His ) at 26°C to an OD 600 of 1.0. The extracts were supplemented with imidazole to a final concentration of 20 mM, incubated at 25 or 37°C for 5 min, as indicated, and subjected to Ni 2+ chelation chromatography using <t>buffer</t> A (see Materials and methods). Equivalent aliquots (2.5, 5 and 10 µl) of the Ni 2+ -NTA–silica eluates were separated by SDS–PAGE using 4–20% gels and subjected to immunoblot analysis using rabbit polyclonal antibodies against the proteins shown on the left, at the following dilutions: PRT1, 1:3000; TIF32, 1:3000; NIP1, 1:1000; TIF34, 1:500; TIF35, 1:5000; eIF5, 1:10000; SUI1/eIF1, 1:1000. Immune complexes were detected by chemiluminescence (ECL™, Amersham Pharmacia Biotech) using horseradish peroxidase-conjugated secondary antibodies (Amersham Pharmacia Biotech).
    Buffer A, supplied by Boehringer Mannheim, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/buffer a/product/Boehringer Mannheim
    Average 86 stars, based on 1 article reviews
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    buffer a - by Bioz Stars, 2021-03
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    86
    Boehringer Mannheim m phenylmethylsulfonyl fluoride
    Fig. 3. Ni 2+ affinity purification of intact eIF3 complexes containing eIF1 and eIF5 from heat-treated extract containing His 8 -prt1-1. Translation extracts were prepared from strains LPY201 ( PRT1-His ) and LPY202 ( prt1-1-His ) at 26°C to an OD 600 of 1.0. The extracts were supplemented with imidazole to a final concentration of 20 mM, incubated at 25 or 37°C for 5 min, as indicated, and subjected to Ni 2+ chelation chromatography using <t>buffer</t> A (see Materials and methods). Equivalent aliquots (2.5, 5 and 10 µl) of the Ni 2+ -NTA–silica eluates were separated by SDS–PAGE using 4–20% gels and subjected to immunoblot analysis using rabbit polyclonal antibodies against the proteins shown on the left, at the following dilutions: PRT1, 1:3000; TIF32, 1:3000; NIP1, 1:1000; TIF34, 1:500; TIF35, 1:5000; eIF5, 1:10000; SUI1/eIF1, 1:1000. Immune complexes were detected by chemiluminescence (ECL™, Amersham Pharmacia Biotech) using horseradish peroxidase-conjugated secondary antibodies (Amersham Pharmacia Biotech).
    M Phenylmethylsulfonyl Fluoride, supplied by Boehringer Mannheim, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/m phenylmethylsulfonyl fluoride/product/Boehringer Mannheim
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    m phenylmethylsulfonyl fluoride - by Bioz Stars, 2021-03
    86/100 stars
      Buy from Supplier

    86
    Boehringer Mannheim phenylmethylsulfonyl fluoride
    Fig. 3. Ni 2+ affinity purification of intact eIF3 complexes containing eIF1 and eIF5 from heat-treated extract containing His 8 -prt1-1. Translation extracts were prepared from strains LPY201 ( PRT1-His ) and LPY202 ( prt1-1-His ) at 26°C to an OD 600 of 1.0. The extracts were supplemented with imidazole to a final concentration of 20 mM, incubated at 25 or 37°C for 5 min, as indicated, and subjected to Ni 2+ chelation chromatography using <t>buffer</t> A (see Materials and methods). Equivalent aliquots (2.5, 5 and 10 µl) of the Ni 2+ -NTA–silica eluates were separated by SDS–PAGE using 4–20% gels and subjected to immunoblot analysis using rabbit polyclonal antibodies against the proteins shown on the left, at the following dilutions: PRT1, 1:3000; TIF32, 1:3000; NIP1, 1:1000; TIF34, 1:500; TIF35, 1:5000; eIF5, 1:10000; SUI1/eIF1, 1:1000. Immune complexes were detected by chemiluminescence (ECL™, Amersham Pharmacia Biotech) using horseradish peroxidase-conjugated secondary antibodies (Amersham Pharmacia Biotech).
    Phenylmethylsulfonyl Fluoride, supplied by Boehringer Mannheim, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/phenylmethylsulfonyl fluoride/product/Boehringer Mannheim
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    phenylmethylsulfonyl fluoride - by Bioz Stars, 2021-03
    86/100 stars
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    Fig. 3. Ni 2+ affinity purification of intact eIF3 complexes containing eIF1 and eIF5 from heat-treated extract containing His 8 -prt1-1. Translation extracts were prepared from strains LPY201 ( PRT1-His ) and LPY202 ( prt1-1-His ) at 26°C to an OD 600 of 1.0. The extracts were supplemented with imidazole to a final concentration of 20 mM, incubated at 25 or 37°C for 5 min, as indicated, and subjected to Ni 2+ chelation chromatography using buffer A (see Materials and methods). Equivalent aliquots (2.5, 5 and 10 µl) of the Ni 2+ -NTA–silica eluates were separated by SDS–PAGE using 4–20% gels and subjected to immunoblot analysis using rabbit polyclonal antibodies against the proteins shown on the left, at the following dilutions: PRT1, 1:3000; TIF32, 1:3000; NIP1, 1:1000; TIF34, 1:500; TIF35, 1:5000; eIF5, 1:10000; SUI1/eIF1, 1:1000. Immune complexes were detected by chemiluminescence (ECL™, Amersham Pharmacia Biotech) using horseradish peroxidase-conjugated secondary antibodies (Amersham Pharmacia Biotech).

    Journal: The EMBO Journal

    Article Title: A subcomplex of three eIF3 subunits binds eIF1 and eIF5 and stimulates ribosome binding of mRNA and tRNAiMet

    doi: 10.1093/emboj/20.11.2954

    Figure Lengend Snippet: Fig. 3. Ni 2+ affinity purification of intact eIF3 complexes containing eIF1 and eIF5 from heat-treated extract containing His 8 -prt1-1. Translation extracts were prepared from strains LPY201 ( PRT1-His ) and LPY202 ( prt1-1-His ) at 26°C to an OD 600 of 1.0. The extracts were supplemented with imidazole to a final concentration of 20 mM, incubated at 25 or 37°C for 5 min, as indicated, and subjected to Ni 2+ chelation chromatography using buffer A (see Materials and methods). Equivalent aliquots (2.5, 5 and 10 µl) of the Ni 2+ -NTA–silica eluates were separated by SDS–PAGE using 4–20% gels and subjected to immunoblot analysis using rabbit polyclonal antibodies against the proteins shown on the left, at the following dilutions: PRT1, 1:3000; TIF32, 1:3000; NIP1, 1:1000; TIF34, 1:500; TIF35, 1:5000; eIF5, 1:10000; SUI1/eIF1, 1:1000. Immune complexes were detected by chemiluminescence (ECL™, Amersham Pharmacia Biotech) using horseradish peroxidase-conjugated secondary antibodies (Amersham Pharmacia Biotech).

    Article Snippet: The cell pellet was resuspended at 1 ml per 1 g of buffer A [20 mM Tris–HCl pH 7.5, 100 mM KCl, 5 mM MgCl2 , 0.5 mM β-mercaptoethanol, 1 mM phenylmethylsulfonyl fluoride (PMSF), 20 mM imidazole, 10% glycerol, 1× complete protease inhibitor cocktail minus EDTA (Boehringer Mannheim)].

    Techniques: Affinity Purification, Concentration Assay, Incubation, Chromatography, SDS Page