complete mini protease inhibitor cocktail  (Roche)

 
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    Structured Review

    Roche complete mini protease inhibitor cocktail
    Expression of conventional breast cancer biomarkers and selected multidrug resistance genes. Cell lysates were prepared in radioimmunoprecipitation assay buffer supplemented with <t>cOmplete</t> Mini protease inhibitor and PhosSTOP phosphatase inhibitor. Quantities (10–50 μg) of total protein were run on a 10 % gel (MDR1), 4–20 % precast gels (epidermal growth factor receptor [EGFR], oestrogen receptor [ER], progesterone receptor [PR], type II topoisomerase [TOPO IIα]) and Any kD Mini-PROTEAN TGX Precast Protein Gels (human epidermal growth factor receptor 2 [HER2], HER3), transferred onto polyvinylidene fluoride membrane and developed using chemiluminescence substrate. Nat native, Epi-R epirubicin-resistant, GAPDH glyceraldehyde-3-phosphate dehydrogenase
    Complete Mini Protease Inhibitor Cocktail, supplied by Roche, used in various techniques. Bioz Stars score: 99/100, based on 350 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    complete mini protease inhibitor cocktail - by Bioz Stars, 2020-04
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    Images

    1) Product Images from "Downregulation of histone H2A and H2B pathways is associated with anthracycline sensitivity in breast cancer"

    Article Title: Downregulation of histone H2A and H2B pathways is associated with anthracycline sensitivity in breast cancer

    Journal: Breast Cancer Research : BCR

    doi: 10.1186/s13058-016-0676-6

    Expression of conventional breast cancer biomarkers and selected multidrug resistance genes. Cell lysates were prepared in radioimmunoprecipitation assay buffer supplemented with cOmplete Mini protease inhibitor and PhosSTOP phosphatase inhibitor. Quantities (10–50 μg) of total protein were run on a 10 % gel (MDR1), 4–20 % precast gels (epidermal growth factor receptor [EGFR], oestrogen receptor [ER], progesterone receptor [PR], type II topoisomerase [TOPO IIα]) and Any kD Mini-PROTEAN TGX Precast Protein Gels (human epidermal growth factor receptor 2 [HER2], HER3), transferred onto polyvinylidene fluoride membrane and developed using chemiluminescence substrate. Nat native, Epi-R epirubicin-resistant, GAPDH glyceraldehyde-3-phosphate dehydrogenase
    Figure Legend Snippet: Expression of conventional breast cancer biomarkers and selected multidrug resistance genes. Cell lysates were prepared in radioimmunoprecipitation assay buffer supplemented with cOmplete Mini protease inhibitor and PhosSTOP phosphatase inhibitor. Quantities (10–50 μg) of total protein were run on a 10 % gel (MDR1), 4–20 % precast gels (epidermal growth factor receptor [EGFR], oestrogen receptor [ER], progesterone receptor [PR], type II topoisomerase [TOPO IIα]) and Any kD Mini-PROTEAN TGX Precast Protein Gels (human epidermal growth factor receptor 2 [HER2], HER3), transferred onto polyvinylidene fluoride membrane and developed using chemiluminescence substrate. Nat native, Epi-R epirubicin-resistant, GAPDH glyceraldehyde-3-phosphate dehydrogenase

    Techniques Used: Expressing, Radio Immunoprecipitation, Protease Inhibitor

    2) Product Images from "Cytotoxic Effects of Tetracycline Analogues (Doxycycline, Minocycline and COL-3) in Acute Myeloid Leukemia HL-60 Cells"

    Article Title: Cytotoxic Effects of Tetracycline Analogues (Doxycycline, Minocycline and COL-3) in Acute Myeloid Leukemia HL-60 Cells

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0114457

    The effect of TCNAs on the viability of human leukemic HL-60 cell line. HL-60 cells were incubated with DOXY, MINO or COL-3 in concentrations within the range 0.5–100 µg/ml for 24 h. Cells incubated in DMSO in a final concentration of 0.2% were used as controls for solvent toxicity and cells incubated in complete medium served as controls. Viability was studied using resazurin fluorescence assay and expressed as percent of the control. Results are presented as a mean ± SD of three independent experiments.
    Figure Legend Snippet: The effect of TCNAs on the viability of human leukemic HL-60 cell line. HL-60 cells were incubated with DOXY, MINO or COL-3 in concentrations within the range 0.5–100 µg/ml for 24 h. Cells incubated in DMSO in a final concentration of 0.2% were used as controls for solvent toxicity and cells incubated in complete medium served as controls. Viability was studied using resazurin fluorescence assay and expressed as percent of the control. Results are presented as a mean ± SD of three independent experiments.

    Techniques Used: Incubation, Concentration Assay, Fluorescence

    TCNAs-mediated poly (ADP-ribose) polymerase-1(PARP-1) cleavage. HL-60 cells were treated with DOXY or MINO in final concentrations of 2.5, 10 and 25 µg/ml or COL-3 in final concentrations of 0.75, 1 and 5 µg/ml for 6 and 24 h. Cells incubated with DMSO in a final concentration of 0.2% served as controls for solvent toxicity, cells incubated with complete medium served as controls. The pattern of PARP-1 cleavage was assessed using WB. Actin was used as a control for equal protein loading. Co: control, D: DOXY, M: MINO and C: COL-3.
    Figure Legend Snippet: TCNAs-mediated poly (ADP-ribose) polymerase-1(PARP-1) cleavage. HL-60 cells were treated with DOXY or MINO in final concentrations of 2.5, 10 and 25 µg/ml or COL-3 in final concentrations of 0.75, 1 and 5 µg/ml for 6 and 24 h. Cells incubated with DMSO in a final concentration of 0.2% served as controls for solvent toxicity, cells incubated with complete medium served as controls. The pattern of PARP-1 cleavage was assessed using WB. Actin was used as a control for equal protein loading. Co: control, D: DOXY, M: MINO and C: COL-3.

    Techniques Used: Incubation, Concentration Assay, Western Blot

    The effect of Z-VAD-FMK on TCNAs-induced cytotoxicity. HL-60 cells were treated with Z-VAD-FMK in a final concentration of 100 µM for 1 h before treatment with TCNAs for 6 h and 24 h. Cells incubated with DMSO in a final concentration of 0.2% served as controls for solvent toxicity, cells incubated in complete medium served as controls. (A) Total cell death was assessed using resazurin viability assay and calculated as 100% - viability %. (B) Apoptosis was assessed by morphological criteria in Giemsa staining. DMSO and Z-VAD-FMK exerted no cytotoxic effects on HL-60 cells compared to controls incubated with complete medium (data not shown on the graph). Results are presented as mean ± SD of three independent experiments. D: DOXY 25 µg/ml, M: MINO 25 µg/ml, Z: Z-VAD-FMK 100 µM; C: COL-3 5 µg/ml in total cell death experiments (A) and 2.5 µg/ml apoptosis experiments (B).
    Figure Legend Snippet: The effect of Z-VAD-FMK on TCNAs-induced cytotoxicity. HL-60 cells were treated with Z-VAD-FMK in a final concentration of 100 µM for 1 h before treatment with TCNAs for 6 h and 24 h. Cells incubated with DMSO in a final concentration of 0.2% served as controls for solvent toxicity, cells incubated in complete medium served as controls. (A) Total cell death was assessed using resazurin viability assay and calculated as 100% - viability %. (B) Apoptosis was assessed by morphological criteria in Giemsa staining. DMSO and Z-VAD-FMK exerted no cytotoxic effects on HL-60 cells compared to controls incubated with complete medium (data not shown on the graph). Results are presented as mean ± SD of three independent experiments. D: DOXY 25 µg/ml, M: MINO 25 µg/ml, Z: Z-VAD-FMK 100 µM; C: COL-3 5 µg/ml in total cell death experiments (A) and 2.5 µg/ml apoptosis experiments (B).

    Techniques Used: Concentration Assay, Incubation, Viability Assay, Staining

    The role of mitochondria in TCNAs-induced apoptosis. Cytosolic translocation of cytochrome c (cyt c) and Bcl-2 cleavage were assessed with Western blot. Cells were incubated with DOXY or MINO in final concentrations of 2.5, 10 and 25 µg/ml or COL-3 in final concentrations of 0.75, 1 and 5 µg/ml until 24 h. DMSO (0.2%) treated cells served as controls for solvent toxicity, etoposide (6 µg/ml) treated cells as positive controls for apoptosis and cells incubated in complete medium as controls. Co: control, D: DOXY, M: MINO, C: COL-3 and E: etoposide.
    Figure Legend Snippet: The role of mitochondria in TCNAs-induced apoptosis. Cytosolic translocation of cytochrome c (cyt c) and Bcl-2 cleavage were assessed with Western blot. Cells were incubated with DOXY or MINO in final concentrations of 2.5, 10 and 25 µg/ml or COL-3 in final concentrations of 0.75, 1 and 5 µg/ml until 24 h. DMSO (0.2%) treated cells served as controls for solvent toxicity, etoposide (6 µg/ml) treated cells as positive controls for apoptosis and cells incubated in complete medium as controls. Co: control, D: DOXY, M: MINO, C: COL-3 and E: etoposide.

    Techniques Used: Translocation Assay, Western Blot, Incubation

    TCNAs-induced apoptosis. HL-60 cells were incubated with DOXY (A) and MINO (B) in concentrations of 5, 10, 25 and 50 µg/ml or COL-3 (C) in concentrations of 0.75, 1, and 5 µg/ml until 48 h. Cells incubated in DMSO in a final concentration of 0.2% were used as controls for solvent toxicity and cells incubated in complete medium served as controls. Cells were stained with annexin V and PI for 15 min at room temperature in the dark. Twenty thousand events were acquired and analyzed using flow cytometry. Subpopulations are expressed as percentages of total events. Results are expressed as a mean ± SD of three independent experiments. (D), (E): Examples on flow cytometric analysis of TCNAs-induced apoptosis using annexin V/PI assay. D: DOXY, M: MINO COL: COL-3 and PI: propidium iodide.
    Figure Legend Snippet: TCNAs-induced apoptosis. HL-60 cells were incubated with DOXY (A) and MINO (B) in concentrations of 5, 10, 25 and 50 µg/ml or COL-3 (C) in concentrations of 0.75, 1, and 5 µg/ml until 48 h. Cells incubated in DMSO in a final concentration of 0.2% were used as controls for solvent toxicity and cells incubated in complete medium served as controls. Cells were stained with annexin V and PI for 15 min at room temperature in the dark. Twenty thousand events were acquired and analyzed using flow cytometry. Subpopulations are expressed as percentages of total events. Results are expressed as a mean ± SD of three independent experiments. (D), (E): Examples on flow cytometric analysis of TCNAs-induced apoptosis using annexin V/PI assay. D: DOXY, M: MINO COL: COL-3 and PI: propidium iodide.

    Techniques Used: Incubation, Concentration Assay, Staining, Flow Cytometry, Cytometry

    The effect of TCNAs on DNA. HL-60 cells were incubated with DOXY or MINO in final concentrations of 25 µg/ml or COL-3 in a final concentration of 5 µg/ml for 10 min and 1, 2 and 4 h. Cells incubated in DMSO in a final concentration of 0.2% were used as controls for solvent toxicity and cells incubated in complete medium served as controls. Cells incubated with etoposide in a final concentration of 6 µg/ml served as positive controls for apoptosis. The TCNAs-induced DNA damage was assessed using SDS-PAGE and immunostaining for the DNA double strand breaks marker γH2AX. Co: control, E: etoposide, D: DOXY, M: MINO and C: COL-3.
    Figure Legend Snippet: The effect of TCNAs on DNA. HL-60 cells were incubated with DOXY or MINO in final concentrations of 25 µg/ml or COL-3 in a final concentration of 5 µg/ml for 10 min and 1, 2 and 4 h. Cells incubated in DMSO in a final concentration of 0.2% were used as controls for solvent toxicity and cells incubated in complete medium served as controls. Cells incubated with etoposide in a final concentration of 6 µg/ml served as positive controls for apoptosis. The TCNAs-induced DNA damage was assessed using SDS-PAGE and immunostaining for the DNA double strand breaks marker γH2AX. Co: control, E: etoposide, D: DOXY, M: MINO and C: COL-3.

    Techniques Used: Incubation, Concentration Assay, SDS Page, Immunostaining, Marker

    The loss of mitochondrial membrane potential (Δψm) in TCNAs-induced apoptosis. Mitochondrial membrane potential was assessed using tetramethylrhodamine methyl ester (TMRM) and flow cytometry. (A) HL-60 cells were incubated with DOXY or MINO in final concentrations of 25 and 50 µg/ml or (B) COL-3 in final concentrations of 0.5, 1, 2.5 and 5 µg/ml for 2, 4, 6 and 24 h. Cells incubated in DMSO in a final concentration of 0.2% were used as controls for solvent toxicity and cells incubated in complete medium served as controls. Results are expressed as mean ± SD of three independent experiments. (C), (D): Example on flow cytometric analysis of loss of Δψm using TMRM assay. D: DOXY, M: MINO, COL: COL-3.
    Figure Legend Snippet: The loss of mitochondrial membrane potential (Δψm) in TCNAs-induced apoptosis. Mitochondrial membrane potential was assessed using tetramethylrhodamine methyl ester (TMRM) and flow cytometry. (A) HL-60 cells were incubated with DOXY or MINO in final concentrations of 25 and 50 µg/ml or (B) COL-3 in final concentrations of 0.5, 1, 2.5 and 5 µg/ml for 2, 4, 6 and 24 h. Cells incubated in DMSO in a final concentration of 0.2% were used as controls for solvent toxicity and cells incubated in complete medium served as controls. Results are expressed as mean ± SD of three independent experiments. (C), (D): Example on flow cytometric analysis of loss of Δψm using TMRM assay. D: DOXY, M: MINO, COL: COL-3.

    Techniques Used: Flow Cytometry, Cytometry, Incubation, Concentration Assay

    3) Product Images from "Evolution of Recombinant Lymphocytic Choriomeningitis Virus/Lassa Virus In Vivo Highlights the Importance of the GPC Cytosolic Tail in Viral Fitness"

    Article Title: Evolution of Recombinant Lymphocytic Choriomeningitis Virus/Lassa Virus In Vivo Highlights the Importance of the GPC Cytosolic Tail in Viral Fitness

    Journal: Journal of Virology

    doi: 10.1128/JVI.00236-14

    Mutation in the GP-2 cytosolic tail increases fitness of virus. (A) The sequence of the quasispecies with an emerging mutation in GP-2 was determined by classical Sanger sequencing of the complete genome. PCR amplicons generated by one-step RT-PCR from
    Figure Legend Snippet: Mutation in the GP-2 cytosolic tail increases fitness of virus. (A) The sequence of the quasispecies with an emerging mutation in GP-2 was determined by classical Sanger sequencing of the complete genome. PCR amplicons generated by one-step RT-PCR from

    Techniques Used: Mutagenesis, Sequencing, Polymerase Chain Reaction, Generated, Reverse Transcription Polymerase Chain Reaction

    4) Product Images from "Knockdown of Laminin gamma-3 (Lamc3) impairs motoneuron guidance in the zebrafish embryo"

    Article Title: Knockdown of Laminin gamma-3 (Lamc3) impairs motoneuron guidance in the zebrafish embryo

    Journal: Wellcome Open Research

    doi: 10.12688/wellcomeopenres.12394.1

    CRISPR/Cas9 mutagenesis of lamc3 causes defects in parachordal chain (PAC) development. ( A ) Design of sgRNAs targeted to exon1 of the lamc3 gene. Each sgRNA targets a restriction endonuclease site used for genotyping and is upstream of a protospacer adjustment motif (PAM). ( B – D ) Cas9/sgRNA injected embryos (as labelled) at 50 hpf. ( B – D ) Brightfield images show no developmental defects or developmental delay. ( B′ – D′ ) Whole embryo images of Tg( fli1a :egfp) fluorescence area indicated by white dotted line is enlarged in ( B′′ – D′′ ) the PAC in each hemisegment is indicated with a red asterisk. ( E ) Quantification of Tg( fli1a :egfp) embryos at 50 hpf with PAC defects. P-values determined by Fisher’s exact Two-tailed test compared to Cas9 alone controls. ( F – I ) 2% agarose genotyping gels with 100 bp ladder. ( F ) Schematic diagram of the region amplified by PCR contains two SphI sites (orange), which would digest into three fragments of 83, 164 and 263 bp in length. Predicted mutation of one of these sites will produce two bands of 83 and 427 bp in length. A natural variant contains a SNP in the second SphI site, complete digestion of this variant would produce two bands of 164 and 346 bp in length. Mutation of the target SphI site would prevent digestion of the fragment. Digestion of the amplicon with Taq α I (blue) in wild type embryos will yield two fragments of 204 and 306 bp in size. ( G ) Restriction endonuclease digest of target region shows undigested mutant products (black asterisk) at a size of 427/510 bp in sgRNA1-injected embryos. ( H ) Fewer undigested mutant products (510 bp) were observed in sgRNA2-injected embryos. ( I ) Brightfield images sgRNA1/Cas9 injected embryo at 5 dpf shows severe oedema. ( J ) Genotyping of 3-month adult fish. A, anterior; b/bp, base pairs; d, days post fertilisation; D, dorsal; kb, kilobases; mpf, months post fertilisation; n, number of embryos; PAC, parachordal chain; PAM, protospacer adjustment motif; SBMO, splice-blocking morpholino; sgRNA, short guide RNA; U, undigested control;.
    Figure Legend Snippet: CRISPR/Cas9 mutagenesis of lamc3 causes defects in parachordal chain (PAC) development. ( A ) Design of sgRNAs targeted to exon1 of the lamc3 gene. Each sgRNA targets a restriction endonuclease site used for genotyping and is upstream of a protospacer adjustment motif (PAM). ( B – D ) Cas9/sgRNA injected embryos (as labelled) at 50 hpf. ( B – D ) Brightfield images show no developmental defects or developmental delay. ( B′ – D′ ) Whole embryo images of Tg( fli1a :egfp) fluorescence area indicated by white dotted line is enlarged in ( B′′ – D′′ ) the PAC in each hemisegment is indicated with a red asterisk. ( E ) Quantification of Tg( fli1a :egfp) embryos at 50 hpf with PAC defects. P-values determined by Fisher’s exact Two-tailed test compared to Cas9 alone controls. ( F – I ) 2% agarose genotyping gels with 100 bp ladder. ( F ) Schematic diagram of the region amplified by PCR contains two SphI sites (orange), which would digest into three fragments of 83, 164 and 263 bp in length. Predicted mutation of one of these sites will produce two bands of 83 and 427 bp in length. A natural variant contains a SNP in the second SphI site, complete digestion of this variant would produce two bands of 164 and 346 bp in length. Mutation of the target SphI site would prevent digestion of the fragment. Digestion of the amplicon with Taq α I (blue) in wild type embryos will yield two fragments of 204 and 306 bp in size. ( G ) Restriction endonuclease digest of target region shows undigested mutant products (black asterisk) at a size of 427/510 bp in sgRNA1-injected embryos. ( H ) Fewer undigested mutant products (510 bp) were observed in sgRNA2-injected embryos. ( I ) Brightfield images sgRNA1/Cas9 injected embryo at 5 dpf shows severe oedema. ( J ) Genotyping of 3-month adult fish. A, anterior; b/bp, base pairs; d, days post fertilisation; D, dorsal; kb, kilobases; mpf, months post fertilisation; n, number of embryos; PAC, parachordal chain; PAM, protospacer adjustment motif; SBMO, splice-blocking morpholino; sgRNA, short guide RNA; U, undigested control;.

    Techniques Used: CRISPR, Mutagenesis, Injection, Fluorescence, Two Tailed Test, Amplification, Polymerase Chain Reaction, Variant Assay, Fluorescence In Situ Hybridization, Blocking Assay

    5) Product Images from "Calcium Influx Rescues Adenylate Cyclase-Hemolysin from Rapid Cell Membrane Removal and Enables Phagocyte Permeabilization by Toxin Pores"

    Article Title: Calcium Influx Rescues Adenylate Cyclase-Hemolysin from Rapid Cell Membrane Removal and Enables Phagocyte Permeabilization by Toxin Pores

    Journal: PLoS Pathogens

    doi: 10.1371/journal.ppat.1002580

    Macropinocytosed dCyaA-KP toxoid is rapidly degraded and its capacity to deliver epitopes for presentation on MHC class II molecules is compromised. (A) 10 6 J774A.1 cells were preincubated for 30 minutes at 37°C in 1 ml of D-MEM medium alone, or in D-MEM containing 1 mM chloroquine plus a cocktail of protease inhibitors (Complete Mini, Roche), or in D-MEM containing 10 mM 2DG and 0.01% of sodium azide, respectively. dCyaA or dCyaA-KP toxoids were added to a final concentration of 1 µg/ml and after continued incubation the cells were washed three times in ice-cold D-MEM at the indicated times and lyzed on ice during 30 minutes in TBS buffer containing 1% Triton X-100 and the protease inhibitor cocktail (Complete Mini, Roche). Cell nuclei were removed by centrifugation at 13,000 RPM for 10 min and supernatants of lyzed cells were separated by 7.5% SDS-PAGE. CyaA fragments were detected in Western blots using the 9D4 antibody recognizing the C-terminal RTX repeats of CyaA. The arrow indicates migration of full-length (undegraded) 200 kDa CyaA. (B) BMDC from C57BL/6 mice were pulsed with indicated concentrations of dCyaA or dCyaA-KP, or (C) with dCyaA-OVA 258–276 or dCyaA-OVA 258–276 -KP and following medium disposal, the MalE-specific hybridoma CRMC3 (B) or OVA 258–276 -specific hybridoma MF2.2D9 cells (C) were added, respectively. Secretion of IL-2 by antigen-stimulated CRMC3 hybridoma (B) was determined as proliferation of the IL-2-dependent CTL-L cell line and the results are expressed in cpm ± SE of duplicate samples. The concentration of IL-2 produced by MF2.2D9 cells upon antigenic stimulation (C) was determined by sandwich ELISA. A representative Western blot from 3 independent experiments and the averaged values from two independent antigen presentation experiments performed in duplicates (n = 4), are shown.
    Figure Legend Snippet: Macropinocytosed dCyaA-KP toxoid is rapidly degraded and its capacity to deliver epitopes for presentation on MHC class II molecules is compromised. (A) 10 6 J774A.1 cells were preincubated for 30 minutes at 37°C in 1 ml of D-MEM medium alone, or in D-MEM containing 1 mM chloroquine plus a cocktail of protease inhibitors (Complete Mini, Roche), or in D-MEM containing 10 mM 2DG and 0.01% of sodium azide, respectively. dCyaA or dCyaA-KP toxoids were added to a final concentration of 1 µg/ml and after continued incubation the cells were washed three times in ice-cold D-MEM at the indicated times and lyzed on ice during 30 minutes in TBS buffer containing 1% Triton X-100 and the protease inhibitor cocktail (Complete Mini, Roche). Cell nuclei were removed by centrifugation at 13,000 RPM for 10 min and supernatants of lyzed cells were separated by 7.5% SDS-PAGE. CyaA fragments were detected in Western blots using the 9D4 antibody recognizing the C-terminal RTX repeats of CyaA. The arrow indicates migration of full-length (undegraded) 200 kDa CyaA. (B) BMDC from C57BL/6 mice were pulsed with indicated concentrations of dCyaA or dCyaA-KP, or (C) with dCyaA-OVA 258–276 or dCyaA-OVA 258–276 -KP and following medium disposal, the MalE-specific hybridoma CRMC3 (B) or OVA 258–276 -specific hybridoma MF2.2D9 cells (C) were added, respectively. Secretion of IL-2 by antigen-stimulated CRMC3 hybridoma (B) was determined as proliferation of the IL-2-dependent CTL-L cell line and the results are expressed in cpm ± SE of duplicate samples. The concentration of IL-2 produced by MF2.2D9 cells upon antigenic stimulation (C) was determined by sandwich ELISA. A representative Western blot from 3 independent experiments and the averaged values from two independent antigen presentation experiments performed in duplicates (n = 4), are shown.

    Techniques Used: Concentration Assay, Incubation, Protease Inhibitor, Centrifugation, SDS Page, Western Blot, Migration, Mouse Assay, CTL Assay, Produced, Sandwich ELISA

    Related Articles

    Diagnostic Assay:

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    Article Snippet: After an overnight fast, blood was collected from the orbital sinus and plasma was separated for measuring IGF-1 with ACTIVE Mouse/Rat IGF-1 RIA kit (Diagnostic Systems Laboratories, Inc.). .. To determine the total content of the protein level of pituitary-specific hormones, the pituitaries from 34-wk-old mice were dissected and lysed in RIPA Lysis buffer [1% Triton X-100, 150 mM NaCl, 10 mM Tris-HCl (pH 7.5), 1 mM EDTA, 1% NP-40, complete mini Protease Inhibitor Cocktail, ROCHE REF. 11836153001] and homogenizied.

    Centrifugation:

    Article Title: Isolation and characterization of the heat shock RNA 1 (HSR1)
    Article Snippet: Add 1 tablet of Roche Complete Mini Protease Inhibitor Cocktail per 1 liter of original culture. .. During the centrifugation prepare GSH-Sepharose as follows: Resuspend the GSH-Sepharose by vigorous shaking of the bottle and transfer 1.33 ml of the suspension for every liter of original bacterial culture into a 15 ml screw-cap tube (Falcon).

    Article Title: Calcium Influx Rescues Adenylate Cyclase-Hemolysin from Rapid Cell Membrane Removal and Enables Phagocyte Permeabilization by Toxin Pores
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    Article Title: Cold-Shock-Domain Protein A (CSDA) Contributes Posttranscriptionally to Gonadotropin-Releasing Hormone-Regulated Expression of Egr1 and Indirectly to Lhb 1
    Article Snippet: .. The αT3 and LβT2 cells were grown to 75% confluence, then treated with 100 nM GnRH and harvested at the indicated time points by washing with cold PBS, then lysing with cold lysis buffer (50 mM Tris [pH 7.5], 200 mM NaCl, 0.5% Igepal, 0.1% SDS, 1 mM EDTA, Complete Mini Protease Inhibitor cocktail [Roche]) on ice for 30 min followed by centrifugation at 16 000 × g for 10 min. .. Protein concentrations were determined using Coomassie Plus protein assay (Pierce).

    Blocking Assay:

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    Real-time Polymerase Chain Reaction:

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    Incubation:

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    Article Snippet: Western blot analysis Cells were collected and then lysed in RIPA lysis and extraction buffer (Thermo Scientific) supplemented with proteinase (Roche, 11836153001) and phosphatase (PhosSTOP™, Sigma, 4906845001). .. Membranes were incubated overnight at 4°C with either MCT2 (LabNed, 0315312; 1:200 dilution), GPT2 (Santa Cruz, 398383; 1:500 dilution), P5CS (Santa Cruz, 515443; 1:500 dilution), GDH (Abcam, 153973; 1:1000 dilution), P4HA1 (Abcam, 59497; 1:2000 dilution), β-Actin (Sigma A5441; 1:10000 dilution) or ERK1/2 (Cell Signaling 4695S; 1:1000 dilution) primary antibodies.

    Article Title: Calcium Influx Rescues Adenylate Cyclase-Hemolysin from Rapid Cell Membrane Removal and Enables Phagocyte Permeabilization by Toxin Pores
    Article Snippet: J774A.1 cells (2×107 ) were washed with prewarmed DMEM and incubated with 500 ng of CyaA-derived proteins at 37°C for 10 min. .. Cells were washed with ice-cold PBS, scraped from the Petri dish and extracted at 4°C in 200 µl of TBS buffer (20 mM Tris-HCl, pH 7.5, 150 mM NaCl) containing 1% Triton X-100, 1 mM EDTA, 10 mM NaF and a Complete Mini protease inhibitor cocktail (Roche) for 60 min.

    Expressing:

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    BIA-KA:

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    Western Blot:

    Article Title: Functional IRF3 deficiency in a patient with herpes simplex encephalitis
    Article Snippet: .. PBMCs used for Western blotting were thawed, washed in PBS, and lysed in a Triton-based lysis buffer (Cell Signaling Technologies) supplemented with Complete-mini protease inhibitor cocktail following the manufacturer’s instructions (Roche). ..

    Article Title: Knockdown of Laminin gamma-3 (Lamc3) impairs motoneuron guidance in the zebrafish embryo
    Article Snippet: Paragraph title: Protein extraction and western blot ... Frozen embryos were thawed and homogenised in 100 μl of protein extraction buffer (1% IGEPAL, 150 mM NaCl, 20 mM Tris pH 7.5, 2 mM EDTA, 50 mM NaF, 1mM sodium pyrophosphate) with 1× cOmplete Mini Protease Inhibitor Cocktail (1183615300, Roche) overnight at 4°C with rotation.

    Article Title: Hypoxia induces TFE3 expression in head and neck squamous cell carcinoma
    Article Snippet: .. Western blot analysis Cultured cells were lysed in T-PER (Pierce, Rockford, IL) containing a complete mini protease inhibitor cocktail and phosphate inhibitors (Roche, Branchburg, NJ). .. Tongue mucosa harvested from two individual Tgfbr1 flox/flox /Pten flox/flox mice and two Tgfbr1/Pten 2cKO mice, and five tumors harvested from Tgfbr1/Pten 2cKO mice, were used for Western blot analysis.

    Article Title: PICK1 Deficiency Impairs Secretory Vesicle Biogenesis and Leads to Growth Retardation and Decreased Glucose TolerancePICK1 and ICA69 Control Insulin Granule Trafficking and Their Deficiencies Lead to Impaired Glucose ToleranceA Pair of Crescent-shaped Proteins Shape Vesicles at the Golgi
    Article Snippet: To determine the total content of the protein level of pituitary-specific hormones, the pituitaries from 34-wk-old mice were dissected and lysed in RIPA Lysis buffer [1% Triton X-100, 150 mM NaCl, 10 mM Tris-HCl (pH 7.5), 1 mM EDTA, 1% NP-40, complete mini Protease Inhibitor Cocktail, ROCHE REF. 11836153001] and homogenizied. .. Insulin receptor expression in liver samples was determined by Western blotting analysis on samples homogenized in the previously mentioned RIPA buffer.

    Article Title: Breast cancer cells rely on environmental pyruvate to shape the metastatic niche
    Article Snippet: .. Western blot analysis Cells were collected and then lysed in RIPA lysis and extraction buffer (Thermo Scientific) supplemented with proteinase (Roche, 11836153001) and phosphatase (PhosSTOP™, Sigma, 4906845001). .. Protein amount was measured using a pierce BCA protein assay kit (Thermo Scientific).

    Article Title: Calcium Influx Rescues Adenylate Cyclase-Hemolysin from Rapid Cell Membrane Removal and Enables Phagocyte Permeabilization by Toxin Pores
    Article Snippet: Paragraph title: Isolation of detergent resistant membrane and western blotting ... Cells were washed with ice-cold PBS, scraped from the Petri dish and extracted at 4°C in 200 µl of TBS buffer (20 mM Tris-HCl, pH 7.5, 150 mM NaCl) containing 1% Triton X-100, 1 mM EDTA, 10 mM NaF and a Complete Mini protease inhibitor cocktail (Roche) for 60 min.

    Article Title: Evolution of Recombinant Lymphocytic Choriomeningitis Virus/Lassa Virus In Vivo Highlights the Importance of the GPC Cytosolic Tail in Viral Fitness
    Article Snippet: Paragraph title: Western blotting. ... Cells were washed twice with cold PBS and lysed in CelLytic M mammalian cell lysis/extraction reagent (Sigma-Aldrich, Switzerland) supplemented with the cOmplete Mini protease inhibitor cocktail (Roche, Switzerland).

    Article Title: Cold-Shock-Domain Protein A (CSDA) Contributes Posttranscriptionally to Gonadotropin-Releasing Hormone-Regulated Expression of Egr1 and Indirectly to Lhb 1
    Article Snippet: Paragraph title: Western Blot ... The αT3 and LβT2 cells were grown to 75% confluence, then treated with 100 nM GnRH and harvested at the indicated time points by washing with cold PBS, then lysing with cold lysis buffer (50 mM Tris [pH 7.5], 200 mM NaCl, 0.5% Igepal, 0.1% SDS, 1 mM EDTA, Complete Mini Protease Inhibitor cocktail [Roche]) on ice for 30 min followed by centrifugation at 16 000 × g for 10 min.

    Transfection:

    Article Title: Isolation and characterization of the heat shock RNA 1 (HSR1)
    Article Snippet: Inoculate a single colony of TOP-10 E. coli cells transfected with GST-HSF1 plasmid (from LB/Amp plate) into 3–5 ml of LB/Amp and grow overnight in the shaker at 37°C. .. Add 1 tablet of Roche Complete Mini Protease Inhibitor Cocktail per 1 liter of original culture.

    Protease Inhibitor:

    Article Title: Isolation and characterization of the heat shock RNA 1 (HSR1)
    Article Snippet: .. Add 1 tablet of Roche Complete Mini Protease Inhibitor Cocktail per 1 liter of original culture. ..

    Article Title: Functional IRF3 deficiency in a patient with herpes simplex encephalitis
    Article Snippet: .. PBMCs used for Western blotting were thawed, washed in PBS, and lysed in a Triton-based lysis buffer (Cell Signaling Technologies) supplemented with Complete-mini protease inhibitor cocktail following the manufacturer’s instructions (Roche). ..

    Article Title: Knockdown of Laminin gamma-3 (Lamc3) impairs motoneuron guidance in the zebrafish embryo
    Article Snippet: .. Frozen embryos were thawed and homogenised in 100 μl of protein extraction buffer (1% IGEPAL, 150 mM NaCl, 20 mM Tris pH 7.5, 2 mM EDTA, 50 mM NaF, 1mM sodium pyrophosphate) with 1× cOmplete Mini Protease Inhibitor Cocktail (1183615300, Roche) overnight at 4°C with rotation. ..

    Article Title: Hypoxia induces TFE3 expression in head and neck squamous cell carcinoma
    Article Snippet: .. Western blot analysis Cultured cells were lysed in T-PER (Pierce, Rockford, IL) containing a complete mini protease inhibitor cocktail and phosphate inhibitors (Roche, Branchburg, NJ). .. Tongue mucosa harvested from two individual Tgfbr1 flox/flox /Pten flox/flox mice and two Tgfbr1/Pten 2cKO mice, and five tumors harvested from Tgfbr1/Pten 2cKO mice, were used for Western blot analysis.

    Article Title: PICK1 Deficiency Impairs Secretory Vesicle Biogenesis and Leads to Growth Retardation and Decreased Glucose TolerancePICK1 and ICA69 Control Insulin Granule Trafficking and Their Deficiencies Lead to Impaired Glucose ToleranceA Pair of Crescent-shaped Proteins Shape Vesicles at the Golgi
    Article Snippet: .. To determine the total content of the protein level of pituitary-specific hormones, the pituitaries from 34-wk-old mice were dissected and lysed in RIPA Lysis buffer [1% Triton X-100, 150 mM NaCl, 10 mM Tris-HCl (pH 7.5), 1 mM EDTA, 1% NP-40, complete mini Protease Inhibitor Cocktail, ROCHE REF. 11836153001] and homogenizied. .. Three different pituitary hormones were measured using ELISA assay: GH (Millipore), prolactin and ACTH (Calbiotech), and TSH (Phoenix Peptides, Burlingame CA).

    Article Title: Cytotoxic Effects of Tetracycline Analogues (Doxycycline, Minocycline and COL-3) in Acute Myeloid Leukemia HL-60 Cells
    Article Snippet: .. The other materials were purchased as follows: Z-VAD-FMK (Bachem AG, Bubendorf, Switzerland); etoposide (Bristol-Myers Squibb, Bromma, Sweden); RPMI 1640 medium, Dulbecco’s phosphate-buffer saline (PBS) and fetal bovine serum (FBS) (Invitrogen AB, Stockholm, Sweden); resazurin (R & D System, Stockholm, Sweden); cOmplete mini protease inhibitor cocktail (Roche AB, Stockholm, Sweden); propidium iodide (Sigma Aldrich) and annexin V (BD, Stockholm, Sweden). ..

    Article Title: Downregulation of histone H2A and H2B pathways is associated with anthracycline sensitivity in breast cancer
    Article Snippet: .. WCL were prepared by adding equal volumes of 2× RIPA buffer supplemented with 25 U of Benzonase Nuclease (Sigma-Aldrich) and cOmplete Mini Protease Inhibitor Cocktail (Roche), incubating on ice for 30 minutes and sonicating for 15 minutes with 30-second on-off intervals. ..

    Article Title: Calcium Influx Rescues Adenylate Cyclase-Hemolysin from Rapid Cell Membrane Removal and Enables Phagocyte Permeabilization by Toxin Pores
    Article Snippet: .. Cells were washed with ice-cold PBS, scraped from the Petri dish and extracted at 4°C in 200 µl of TBS buffer (20 mM Tris-HCl, pH 7.5, 150 mM NaCl) containing 1% Triton X-100, 1 mM EDTA, 10 mM NaF and a Complete Mini protease inhibitor cocktail (Roche) for 60 min. .. The lyzate was clarified by centrifugation at 250× g for 5 min and the post-nuclear supernatant was mixed with equal volume of 90% sucrose in TBS.

    Article Title: Evolution of Recombinant Lymphocytic Choriomeningitis Virus/Lassa Virus In Vivo Highlights the Importance of the GPC Cytosolic Tail in Viral Fitness
    Article Snippet: .. Cells were washed twice with cold PBS and lysed in CelLytic M mammalian cell lysis/extraction reagent (Sigma-Aldrich, Switzerland) supplemented with the cOmplete Mini protease inhibitor cocktail (Roche, Switzerland). .. Samples were cleared of cellular debris by centrifugation, mixed in a 1:1 ratio with 2× SDS-PAGE sample buffer (62.5 mM Tris-HCl [pH 6.8], 20% glycerol, 2% SDS, 10% dithiothreitol), boiled for 10 min at 95°C, and centrifuged for 5 min at 13,000 rpm prior to loading.

    Article Title: Cold-Shock-Domain Protein A (CSDA) Contributes Posttranscriptionally to Gonadotropin-Releasing Hormone-Regulated Expression of Egr1 and Indirectly to Lhb 1
    Article Snippet: .. The αT3 and LβT2 cells were grown to 75% confluence, then treated with 100 nM GnRH and harvested at the indicated time points by washing with cold PBS, then lysing with cold lysis buffer (50 mM Tris [pH 7.5], 200 mM NaCl, 0.5% Igepal, 0.1% SDS, 1 mM EDTA, Complete Mini Protease Inhibitor cocktail [Roche]) on ice for 30 min followed by centrifugation at 16 000 × g for 10 min. .. Protein concentrations were determined using Coomassie Plus protein assay (Pierce).

    Transferring:

    Article Title: Knockdown of Laminin gamma-3 (Lamc3) impairs motoneuron guidance in the zebrafish embryo
    Article Snippet: Embryos were de-yolked by pipetting using a 200 μl pipette tip. .. Frozen embryos were thawed and homogenised in 100 μl of protein extraction buffer (1% IGEPAL, 150 mM NaCl, 20 mM Tris pH 7.5, 2 mM EDTA, 50 mM NaF, 1mM sodium pyrophosphate) with 1× cOmplete Mini Protease Inhibitor Cocktail (1183615300, Roche) overnight at 4°C with rotation.

    Cell Culture:

    Article Title: Hypoxia induces TFE3 expression in head and neck squamous cell carcinoma
    Article Snippet: .. Western blot analysis Cultured cells were lysed in T-PER (Pierce, Rockford, IL) containing a complete mini protease inhibitor cocktail and phosphate inhibitors (Roche, Branchburg, NJ). .. Tongue mucosa harvested from two individual Tgfbr1 flox/flox /Pten flox/flox mice and two Tgfbr1/Pten 2cKO mice, and five tumors harvested from Tgfbr1/Pten 2cKO mice, were used for Western blot analysis.

    other:

    Article Title: Calcium Influx Rescues Adenylate Cyclase-Hemolysin from Rapid Cell Membrane Removal and Enables Phagocyte Permeabilization by Toxin Pores
    Article Snippet: Complete Mini protease inhibitors cocktail was purchased from Roche (Basel, Switzerland).

    Imaging:

    Article Title: Downregulation of histone H2A and H2B pathways is associated with anthracycline sensitivity in breast cancer
    Article Snippet: WCL were prepared by adding equal volumes of 2× RIPA buffer supplemented with 25 U of Benzonase Nuclease (Sigma-Aldrich) and cOmplete Mini Protease Inhibitor Cocktail (Roche), incubating on ice for 30 minutes and sonicating for 15 minutes with 30-second on-off intervals. .. Signals were developed with BM Chemiluminescence Blotting Substrate (POD) (Roche) and a ChemiDoc imaging system (Bio-Rad Laboratories).

    Injection:

    Article Title: PICK1 Deficiency Impairs Secretory Vesicle Biogenesis and Leads to Growth Retardation and Decreased Glucose TolerancePICK1 and ICA69 Control Insulin Granule Trafficking and Their Deficiencies Lead to Impaired Glucose ToleranceA Pair of Crescent-shaped Proteins Shape Vesicles at the Golgi
    Article Snippet: Blood was sampled from orbital sinus before and 5 min after injection . .. To determine the total content of the protein level of pituitary-specific hormones, the pituitaries from 34-wk-old mice were dissected and lysed in RIPA Lysis buffer [1% Triton X-100, 150 mM NaCl, 10 mM Tris-HCl (pH 7.5), 1 mM EDTA, 1% NP-40, complete mini Protease Inhibitor Cocktail, ROCHE REF. 11836153001] and homogenizied.

    Recombinant:

    Article Title: Isolation and characterization of the heat shock RNA 1 (HSR1)
    Article Snippet: Paragraph title: 3.1. Purification of recombinant HSF ... Add 1 tablet of Roche Complete Mini Protease Inhibitor Cocktail per 1 liter of original culture.

    Isolation:

    Article Title: Calcium Influx Rescues Adenylate Cyclase-Hemolysin from Rapid Cell Membrane Removal and Enables Phagocyte Permeabilization by Toxin Pores
    Article Snippet: Paragraph title: Isolation of detergent resistant membrane and western blotting ... Cells were washed with ice-cold PBS, scraped from the Petri dish and extracted at 4°C in 200 µl of TBS buffer (20 mM Tris-HCl, pH 7.5, 150 mM NaCl) containing 1% Triton X-100, 1 mM EDTA, 10 mM NaF and a Complete Mini protease inhibitor cocktail (Roche) for 60 min.

    Purification:

    Article Title: Isolation and characterization of the heat shock RNA 1 (HSR1)
    Article Snippet: Paragraph title: 3.1. Purification of recombinant HSF ... Add 1 tablet of Roche Complete Mini Protease Inhibitor Cocktail per 1 liter of original culture.

    Protein Extraction:

    Article Title: Knockdown of Laminin gamma-3 (Lamc3) impairs motoneuron guidance in the zebrafish embryo
    Article Snippet: .. Frozen embryos were thawed and homogenised in 100 μl of protein extraction buffer (1% IGEPAL, 150 mM NaCl, 20 mM Tris pH 7.5, 2 mM EDTA, 50 mM NaF, 1mM sodium pyrophosphate) with 1× cOmplete Mini Protease Inhibitor Cocktail (1183615300, Roche) overnight at 4°C with rotation. ..

    Mouse Assay:

    Article Title: Hypoxia induces TFE3 expression in head and neck squamous cell carcinoma
    Article Snippet: Western blot analysis Cultured cells were lysed in T-PER (Pierce, Rockford, IL) containing a complete mini protease inhibitor cocktail and phosphate inhibitors (Roche, Branchburg, NJ). .. Tongue mucosa harvested from two individual Tgfbr1 flox/flox /Pten flox/flox mice and two Tgfbr1/Pten 2cKO mice, and five tumors harvested from Tgfbr1/Pten 2cKO mice, were used for Western blot analysis.

    Article Title: PICK1 Deficiency Impairs Secretory Vesicle Biogenesis and Leads to Growth Retardation and Decreased Glucose TolerancePICK1 and ICA69 Control Insulin Granule Trafficking and Their Deficiencies Lead to Impaired Glucose ToleranceA Pair of Crescent-shaped Proteins Shape Vesicles at the Golgi
    Article Snippet: .. To determine the total content of the protein level of pituitary-specific hormones, the pituitaries from 34-wk-old mice were dissected and lysed in RIPA Lysis buffer [1% Triton X-100, 150 mM NaCl, 10 mM Tris-HCl (pH 7.5), 1 mM EDTA, 1% NP-40, complete mini Protease Inhibitor Cocktail, ROCHE REF. 11836153001] and homogenizied. .. Three different pituitary hormones were measured using ELISA assay: GH (Millipore), prolactin and ACTH (Calbiotech), and TSH (Phoenix Peptides, Burlingame CA).

    SDS Page:

    Article Title: Evolution of Recombinant Lymphocytic Choriomeningitis Virus/Lassa Virus In Vivo Highlights the Importance of the GPC Cytosolic Tail in Viral Fitness
    Article Snippet: Cells were washed twice with cold PBS and lysed in CelLytic M mammalian cell lysis/extraction reagent (Sigma-Aldrich, Switzerland) supplemented with the cOmplete Mini protease inhibitor cocktail (Roche, Switzerland). .. Samples were cleared of cellular debris by centrifugation, mixed in a 1:1 ratio with 2× SDS-PAGE sample buffer (62.5 mM Tris-HCl [pH 6.8], 20% glycerol, 2% SDS, 10% dithiothreitol), boiled for 10 min at 95°C, and centrifuged for 5 min at 13,000 rpm prior to loading.

    Plasmid Preparation:

    Article Title: Isolation and characterization of the heat shock RNA 1 (HSR1)
    Article Snippet: Inoculate a single colony of TOP-10 E. coli cells transfected with GST-HSF1 plasmid (from LB/Amp plate) into 3–5 ml of LB/Amp and grow overnight in the shaker at 37°C. .. Add 1 tablet of Roche Complete Mini Protease Inhibitor Cocktail per 1 liter of original culture.

    Enzyme-linked Immunosorbent Assay:

    Article Title: PICK1 Deficiency Impairs Secretory Vesicle Biogenesis and Leads to Growth Retardation and Decreased Glucose TolerancePICK1 and ICA69 Control Insulin Granule Trafficking and Their Deficiencies Lead to Impaired Glucose ToleranceA Pair of Crescent-shaped Proteins Shape Vesicles at the Golgi
    Article Snippet: Plasma GH concentration was measured using Rat/Mouse GH ELISA KIT (Millipore) and plasma triglyceride (DT60II Model, Orthoclinical Diagnostics; Sollentuna, Sweden). .. To determine the total content of the protein level of pituitary-specific hormones, the pituitaries from 34-wk-old mice were dissected and lysed in RIPA Lysis buffer [1% Triton X-100, 150 mM NaCl, 10 mM Tris-HCl (pH 7.5), 1 mM EDTA, 1% NP-40, complete mini Protease Inhibitor Cocktail, ROCHE REF. 11836153001] and homogenizied.

    Radio Immunoprecipitation:

    Article Title: Downregulation of histone H2A and H2B pathways is associated with anthracycline sensitivity in breast cancer
    Article Snippet: Immunoblotting Whole-cell lysates (WCL) were prepared in radioimmunoprecipitation assay (RIPA) buffer supplemented with cOmplete Mini Protease and PhosSTOP phosphatase inhibitors (Roche, Laval, QC, Canada). .. WCL were prepared by adding equal volumes of 2× RIPA buffer supplemented with 25 U of Benzonase Nuclease (Sigma-Aldrich) and cOmplete Mini Protease Inhibitor Cocktail (Roche), incubating on ice for 30 minutes and sonicating for 15 minutes with 30-second on-off intervals.

    Concentration Assay:

    Article Title: Isolation and characterization of the heat shock RNA 1 (HSR1)
    Article Snippet: Add IPTG to a final concentration of 0.1 mM and return the culture to the 25–28°C shaker to allow GST-HSF1 synthesis to proceed for 4–6 hours. .. Add 1 tablet of Roche Complete Mini Protease Inhibitor Cocktail per 1 liter of original culture.

    Article Title: PICK1 Deficiency Impairs Secretory Vesicle Biogenesis and Leads to Growth Retardation and Decreased Glucose TolerancePICK1 and ICA69 Control Insulin Granule Trafficking and Their Deficiencies Lead to Impaired Glucose ToleranceA Pair of Crescent-shaped Proteins Shape Vesicles at the Golgi
    Article Snippet: Plasma GH concentration was measured using Rat/Mouse GH ELISA KIT (Millipore) and plasma triglyceride (DT60II Model, Orthoclinical Diagnostics; Sollentuna, Sweden). .. To determine the total content of the protein level of pituitary-specific hormones, the pituitaries from 34-wk-old mice were dissected and lysed in RIPA Lysis buffer [1% Triton X-100, 150 mM NaCl, 10 mM Tris-HCl (pH 7.5), 1 mM EDTA, 1% NP-40, complete mini Protease Inhibitor Cocktail, ROCHE REF. 11836153001] and homogenizied.

    Article Title: Cytotoxic Effects of Tetracycline Analogues (Doxycycline, Minocycline and COL-3) in Acute Myeloid Leukemia HL-60 Cells
    Article Snippet: Stock solutions of DOXY and MINO were prepared in distilled water (dH2 O) in a concentration of 5 mg/ml and stocks of COL-3 in dimethyl sulfoxide (DMSO; Sigma Aldrich, Stockholm, Sweden) in a concentration of 50 mg/ml and stored at −20°C. .. The other materials were purchased as follows: Z-VAD-FMK (Bachem AG, Bubendorf, Switzerland); etoposide (Bristol-Myers Squibb, Bromma, Sweden); RPMI 1640 medium, Dulbecco’s phosphate-buffer saline (PBS) and fetal bovine serum (FBS) (Invitrogen AB, Stockholm, Sweden); resazurin (R & D System, Stockholm, Sweden); cOmplete mini protease inhibitor cocktail (Roche AB, Stockholm, Sweden); propidium iodide (Sigma Aldrich) and annexin V (BD, Stockholm, Sweden).

    Lysis:

    Article Title: Isolation and characterization of the heat shock RNA 1 (HSR1)
    Article Snippet: Resuspend the bacteria in 20 ml of the lysis buffer per 1 liter of original culture. .. Add 1 tablet of Roche Complete Mini Protease Inhibitor Cocktail per 1 liter of original culture.

    Article Title: Functional IRF3 deficiency in a patient with herpes simplex encephalitis
    Article Snippet: .. PBMCs used for Western blotting were thawed, washed in PBS, and lysed in a Triton-based lysis buffer (Cell Signaling Technologies) supplemented with Complete-mini protease inhibitor cocktail following the manufacturer’s instructions (Roche). ..

    Article Title: PICK1 Deficiency Impairs Secretory Vesicle Biogenesis and Leads to Growth Retardation and Decreased Glucose TolerancePICK1 and ICA69 Control Insulin Granule Trafficking and Their Deficiencies Lead to Impaired Glucose ToleranceA Pair of Crescent-shaped Proteins Shape Vesicles at the Golgi
    Article Snippet: .. To determine the total content of the protein level of pituitary-specific hormones, the pituitaries from 34-wk-old mice were dissected and lysed in RIPA Lysis buffer [1% Triton X-100, 150 mM NaCl, 10 mM Tris-HCl (pH 7.5), 1 mM EDTA, 1% NP-40, complete mini Protease Inhibitor Cocktail, ROCHE REF. 11836153001] and homogenizied. .. Three different pituitary hormones were measured using ELISA assay: GH (Millipore), prolactin and ACTH (Calbiotech), and TSH (Phoenix Peptides, Burlingame CA).

    Article Title: Breast cancer cells rely on environmental pyruvate to shape the metastatic niche
    Article Snippet: .. Western blot analysis Cells were collected and then lysed in RIPA lysis and extraction buffer (Thermo Scientific) supplemented with proteinase (Roche, 11836153001) and phosphatase (PhosSTOP™, Sigma, 4906845001). .. Protein amount was measured using a pierce BCA protein assay kit (Thermo Scientific).

    Article Title: Evolution of Recombinant Lymphocytic Choriomeningitis Virus/Lassa Virus In Vivo Highlights the Importance of the GPC Cytosolic Tail in Viral Fitness
    Article Snippet: .. Cells were washed twice with cold PBS and lysed in CelLytic M mammalian cell lysis/extraction reagent (Sigma-Aldrich, Switzerland) supplemented with the cOmplete Mini protease inhibitor cocktail (Roche, Switzerland). .. Samples were cleared of cellular debris by centrifugation, mixed in a 1:1 ratio with 2× SDS-PAGE sample buffer (62.5 mM Tris-HCl [pH 6.8], 20% glycerol, 2% SDS, 10% dithiothreitol), boiled for 10 min at 95°C, and centrifuged for 5 min at 13,000 rpm prior to loading.

    Article Title: Cold-Shock-Domain Protein A (CSDA) Contributes Posttranscriptionally to Gonadotropin-Releasing Hormone-Regulated Expression of Egr1 and Indirectly to Lhb 1
    Article Snippet: .. The αT3 and LβT2 cells were grown to 75% confluence, then treated with 100 nM GnRH and harvested at the indicated time points by washing with cold PBS, then lysing with cold lysis buffer (50 mM Tris [pH 7.5], 200 mM NaCl, 0.5% Igepal, 0.1% SDS, 1 mM EDTA, Complete Mini Protease Inhibitor cocktail [Roche]) on ice for 30 min followed by centrifugation at 16 000 × g for 10 min. .. Protein concentrations were determined using Coomassie Plus protein assay (Pierce).

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  • 99
    Roche complete mini protease inhibitor cocktail
    Expression of conventional breast cancer biomarkers and selected multidrug resistance genes. Cell lysates were prepared in radioimmunoprecipitation assay buffer supplemented with <t>cOmplete</t> Mini protease inhibitor and PhosSTOP phosphatase inhibitor. Quantities (10–50 μg) of total protein were run on a 10 % gel (MDR1), 4–20 % precast gels (epidermal growth factor receptor [EGFR], oestrogen receptor [ER], progesterone receptor [PR], type II topoisomerase [TOPO IIα]) and Any kD Mini-PROTEAN TGX Precast Protein Gels (human epidermal growth factor receptor 2 [HER2], HER3), transferred onto polyvinylidene fluoride membrane and developed using chemiluminescence substrate. Nat native, Epi-R epirubicin-resistant, GAPDH glyceraldehyde-3-phosphate dehydrogenase
    Complete Mini Protease Inhibitor Cocktail, supplied by Roche, used in various techniques. Bioz Stars score: 99/100, based on 351 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/complete mini protease inhibitor cocktail/product/Roche
    Average 99 stars, based on 351 article reviews
    Price from $9.99 to $1999.99
    complete mini protease inhibitor cocktail - by Bioz Stars, 2020-04
    99/100 stars
      Buy from Supplier

    99
    Roche complete mini edta free protease inhibitor cocktail
    Apocatalase polypeptide proteolysis is induced in late exponential growth phase E. faecalis cells. A , lysates from strain OG1RF cells grown in heme-free medium until early exponential growth phase ( sample A ) or late exponential growth phase ( sample B ) and lysates from cells grown in heme-supplemented medium until late exponential growth phase ( sample C ) were analyzed individually or as mixtures as indicated. The samples were incubated at 37 °C, and the presence of KatA polypeptide in aliquots removed at the indicated time points was determined by immuno-blot. B , proteolysis of KatA in sample A mixed with sample B in the presence of 1 m m <t>PMSF</t> and/or <t>EDTA</t> or Complete EDTA-free protease inhibitor cocktail. The reference, sample A + B without any additions, is indicated by Ctrl .
    Complete Mini Edta Free Protease Inhibitor Cocktail, supplied by Roche, used in various techniques. Bioz Stars score: 99/100, based on 17 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/complete mini edta free protease inhibitor cocktail/product/Roche
    Average 99 stars, based on 17 article reviews
    Price from $9.99 to $1999.99
    complete mini edta free protease inhibitor cocktail - by Bioz Stars, 2020-04
    99/100 stars
      Buy from Supplier

    Image Search Results


    Expression of conventional breast cancer biomarkers and selected multidrug resistance genes. Cell lysates were prepared in radioimmunoprecipitation assay buffer supplemented with cOmplete Mini protease inhibitor and PhosSTOP phosphatase inhibitor. Quantities (10–50 μg) of total protein were run on a 10 % gel (MDR1), 4–20 % precast gels (epidermal growth factor receptor [EGFR], oestrogen receptor [ER], progesterone receptor [PR], type II topoisomerase [TOPO IIα]) and Any kD Mini-PROTEAN TGX Precast Protein Gels (human epidermal growth factor receptor 2 [HER2], HER3), transferred onto polyvinylidene fluoride membrane and developed using chemiluminescence substrate. Nat native, Epi-R epirubicin-resistant, GAPDH glyceraldehyde-3-phosphate dehydrogenase

    Journal: Breast Cancer Research : BCR

    Article Title: Downregulation of histone H2A and H2B pathways is associated with anthracycline sensitivity in breast cancer

    doi: 10.1186/s13058-016-0676-6

    Figure Lengend Snippet: Expression of conventional breast cancer biomarkers and selected multidrug resistance genes. Cell lysates were prepared in radioimmunoprecipitation assay buffer supplemented with cOmplete Mini protease inhibitor and PhosSTOP phosphatase inhibitor. Quantities (10–50 μg) of total protein were run on a 10 % gel (MDR1), 4–20 % precast gels (epidermal growth factor receptor [EGFR], oestrogen receptor [ER], progesterone receptor [PR], type II topoisomerase [TOPO IIα]) and Any kD Mini-PROTEAN TGX Precast Protein Gels (human epidermal growth factor receptor 2 [HER2], HER3), transferred onto polyvinylidene fluoride membrane and developed using chemiluminescence substrate. Nat native, Epi-R epirubicin-resistant, GAPDH glyceraldehyde-3-phosphate dehydrogenase

    Article Snippet: WCL were prepared by adding equal volumes of 2× RIPA buffer supplemented with 25 U of Benzonase Nuclease (Sigma-Aldrich) and cOmplete Mini Protease Inhibitor Cocktail (Roche), incubating on ice for 30 minutes and sonicating for 15 minutes with 30-second on-off intervals.

    Techniques: Expressing, Radio Immunoprecipitation, Protease Inhibitor

    Giardia duodenalis -induced increase of epithelial sugar uptake is mediated by proteases. (A) Addition of protease inhibitor cocktails, complete mini (CM) decreased G. duodenalis -enhanced sugar uptake. CM alone (CM control) had no effect on baseline enterocytic

    Journal:

    Article Title: SGLT-1-mediated glucose uptake protects human intestinal epithelial cells against Giardia duodenalis-induced apoptosis

    doi: 10.1016/j.ijpara.2007.12.004

    Figure Lengend Snippet: Giardia duodenalis -induced increase of epithelial sugar uptake is mediated by proteases. (A) Addition of protease inhibitor cocktails, complete mini (CM) decreased G. duodenalis -enhanced sugar uptake. CM alone (CM control) had no effect on baseline enterocytic

    Article Snippet: The protease inhibitor cocktail complete mini® which inhibits serine, cysteine, metalloproteases and calpains, the selective cysteine protease inhibitor E64, and the selective serine protease inhibitor Pefabloc SC were purchased from Roche, Laval, Que., Canada.

    Techniques: Protease Inhibitor

    The effect of TCNAs on the viability of human leukemic HL-60 cell line. HL-60 cells were incubated with DOXY, MINO or COL-3 in concentrations within the range 0.5–100 µg/ml for 24 h. Cells incubated in DMSO in a final concentration of 0.2% were used as controls for solvent toxicity and cells incubated in complete medium served as controls. Viability was studied using resazurin fluorescence assay and expressed as percent of the control. Results are presented as a mean ± SD of three independent experiments.

    Journal: PLoS ONE

    Article Title: Cytotoxic Effects of Tetracycline Analogues (Doxycycline, Minocycline and COL-3) in Acute Myeloid Leukemia HL-60 Cells

    doi: 10.1371/journal.pone.0114457

    Figure Lengend Snippet: The effect of TCNAs on the viability of human leukemic HL-60 cell line. HL-60 cells were incubated with DOXY, MINO or COL-3 in concentrations within the range 0.5–100 µg/ml for 24 h. Cells incubated in DMSO in a final concentration of 0.2% were used as controls for solvent toxicity and cells incubated in complete medium served as controls. Viability was studied using resazurin fluorescence assay and expressed as percent of the control. Results are presented as a mean ± SD of three independent experiments.

    Article Snippet: The other materials were purchased as follows: Z-VAD-FMK (Bachem AG, Bubendorf, Switzerland); etoposide (Bristol-Myers Squibb, Bromma, Sweden); RPMI 1640 medium, Dulbecco’s phosphate-buffer saline (PBS) and fetal bovine serum (FBS) (Invitrogen AB, Stockholm, Sweden); resazurin (R & D System, Stockholm, Sweden); cOmplete mini protease inhibitor cocktail (Roche AB, Stockholm, Sweden); propidium iodide (Sigma Aldrich) and annexin V (BD, Stockholm, Sweden).

    Techniques: Incubation, Concentration Assay, Fluorescence

    TCNAs-mediated poly (ADP-ribose) polymerase-1(PARP-1) cleavage. HL-60 cells were treated with DOXY or MINO in final concentrations of 2.5, 10 and 25 µg/ml or COL-3 in final concentrations of 0.75, 1 and 5 µg/ml for 6 and 24 h. Cells incubated with DMSO in a final concentration of 0.2% served as controls for solvent toxicity, cells incubated with complete medium served as controls. The pattern of PARP-1 cleavage was assessed using WB. Actin was used as a control for equal protein loading. Co: control, D: DOXY, M: MINO and C: COL-3.

    Journal: PLoS ONE

    Article Title: Cytotoxic Effects of Tetracycline Analogues (Doxycycline, Minocycline and COL-3) in Acute Myeloid Leukemia HL-60 Cells

    doi: 10.1371/journal.pone.0114457

    Figure Lengend Snippet: TCNAs-mediated poly (ADP-ribose) polymerase-1(PARP-1) cleavage. HL-60 cells were treated with DOXY or MINO in final concentrations of 2.5, 10 and 25 µg/ml or COL-3 in final concentrations of 0.75, 1 and 5 µg/ml for 6 and 24 h. Cells incubated with DMSO in a final concentration of 0.2% served as controls for solvent toxicity, cells incubated with complete medium served as controls. The pattern of PARP-1 cleavage was assessed using WB. Actin was used as a control for equal protein loading. Co: control, D: DOXY, M: MINO and C: COL-3.

    Article Snippet: The other materials were purchased as follows: Z-VAD-FMK (Bachem AG, Bubendorf, Switzerland); etoposide (Bristol-Myers Squibb, Bromma, Sweden); RPMI 1640 medium, Dulbecco’s phosphate-buffer saline (PBS) and fetal bovine serum (FBS) (Invitrogen AB, Stockholm, Sweden); resazurin (R & D System, Stockholm, Sweden); cOmplete mini protease inhibitor cocktail (Roche AB, Stockholm, Sweden); propidium iodide (Sigma Aldrich) and annexin V (BD, Stockholm, Sweden).

    Techniques: Incubation, Concentration Assay, Western Blot

    The effect of Z-VAD-FMK on TCNAs-induced cytotoxicity. HL-60 cells were treated with Z-VAD-FMK in a final concentration of 100 µM for 1 h before treatment with TCNAs for 6 h and 24 h. Cells incubated with DMSO in a final concentration of 0.2% served as controls for solvent toxicity, cells incubated in complete medium served as controls. (A) Total cell death was assessed using resazurin viability assay and calculated as 100% - viability %. (B) Apoptosis was assessed by morphological criteria in Giemsa staining. DMSO and Z-VAD-FMK exerted no cytotoxic effects on HL-60 cells compared to controls incubated with complete medium (data not shown on the graph). Results are presented as mean ± SD of three independent experiments. D: DOXY 25 µg/ml, M: MINO 25 µg/ml, Z: Z-VAD-FMK 100 µM; C: COL-3 5 µg/ml in total cell death experiments (A) and 2.5 µg/ml apoptosis experiments (B).

    Journal: PLoS ONE

    Article Title: Cytotoxic Effects of Tetracycline Analogues (Doxycycline, Minocycline and COL-3) in Acute Myeloid Leukemia HL-60 Cells

    doi: 10.1371/journal.pone.0114457

    Figure Lengend Snippet: The effect of Z-VAD-FMK on TCNAs-induced cytotoxicity. HL-60 cells were treated with Z-VAD-FMK in a final concentration of 100 µM for 1 h before treatment with TCNAs for 6 h and 24 h. Cells incubated with DMSO in a final concentration of 0.2% served as controls for solvent toxicity, cells incubated in complete medium served as controls. (A) Total cell death was assessed using resazurin viability assay and calculated as 100% - viability %. (B) Apoptosis was assessed by morphological criteria in Giemsa staining. DMSO and Z-VAD-FMK exerted no cytotoxic effects on HL-60 cells compared to controls incubated with complete medium (data not shown on the graph). Results are presented as mean ± SD of three independent experiments. D: DOXY 25 µg/ml, M: MINO 25 µg/ml, Z: Z-VAD-FMK 100 µM; C: COL-3 5 µg/ml in total cell death experiments (A) and 2.5 µg/ml apoptosis experiments (B).

    Article Snippet: The other materials were purchased as follows: Z-VAD-FMK (Bachem AG, Bubendorf, Switzerland); etoposide (Bristol-Myers Squibb, Bromma, Sweden); RPMI 1640 medium, Dulbecco’s phosphate-buffer saline (PBS) and fetal bovine serum (FBS) (Invitrogen AB, Stockholm, Sweden); resazurin (R & D System, Stockholm, Sweden); cOmplete mini protease inhibitor cocktail (Roche AB, Stockholm, Sweden); propidium iodide (Sigma Aldrich) and annexin V (BD, Stockholm, Sweden).

    Techniques: Concentration Assay, Incubation, Viability Assay, Staining

    The role of mitochondria in TCNAs-induced apoptosis. Cytosolic translocation of cytochrome c (cyt c) and Bcl-2 cleavage were assessed with Western blot. Cells were incubated with DOXY or MINO in final concentrations of 2.5, 10 and 25 µg/ml or COL-3 in final concentrations of 0.75, 1 and 5 µg/ml until 24 h. DMSO (0.2%) treated cells served as controls for solvent toxicity, etoposide (6 µg/ml) treated cells as positive controls for apoptosis and cells incubated in complete medium as controls. Co: control, D: DOXY, M: MINO, C: COL-3 and E: etoposide.

    Journal: PLoS ONE

    Article Title: Cytotoxic Effects of Tetracycline Analogues (Doxycycline, Minocycline and COL-3) in Acute Myeloid Leukemia HL-60 Cells

    doi: 10.1371/journal.pone.0114457

    Figure Lengend Snippet: The role of mitochondria in TCNAs-induced apoptosis. Cytosolic translocation of cytochrome c (cyt c) and Bcl-2 cleavage were assessed with Western blot. Cells were incubated with DOXY or MINO in final concentrations of 2.5, 10 and 25 µg/ml or COL-3 in final concentrations of 0.75, 1 and 5 µg/ml until 24 h. DMSO (0.2%) treated cells served as controls for solvent toxicity, etoposide (6 µg/ml) treated cells as positive controls for apoptosis and cells incubated in complete medium as controls. Co: control, D: DOXY, M: MINO, C: COL-3 and E: etoposide.

    Article Snippet: The other materials were purchased as follows: Z-VAD-FMK (Bachem AG, Bubendorf, Switzerland); etoposide (Bristol-Myers Squibb, Bromma, Sweden); RPMI 1640 medium, Dulbecco’s phosphate-buffer saline (PBS) and fetal bovine serum (FBS) (Invitrogen AB, Stockholm, Sweden); resazurin (R & D System, Stockholm, Sweden); cOmplete mini protease inhibitor cocktail (Roche AB, Stockholm, Sweden); propidium iodide (Sigma Aldrich) and annexin V (BD, Stockholm, Sweden).

    Techniques: Translocation Assay, Western Blot, Incubation

    TCNAs-induced apoptosis. HL-60 cells were incubated with DOXY (A) and MINO (B) in concentrations of 5, 10, 25 and 50 µg/ml or COL-3 (C) in concentrations of 0.75, 1, and 5 µg/ml until 48 h. Cells incubated in DMSO in a final concentration of 0.2% were used as controls for solvent toxicity and cells incubated in complete medium served as controls. Cells were stained with annexin V and PI for 15 min at room temperature in the dark. Twenty thousand events were acquired and analyzed using flow cytometry. Subpopulations are expressed as percentages of total events. Results are expressed as a mean ± SD of three independent experiments. (D), (E): Examples on flow cytometric analysis of TCNAs-induced apoptosis using annexin V/PI assay. D: DOXY, M: MINO COL: COL-3 and PI: propidium iodide.

    Journal: PLoS ONE

    Article Title: Cytotoxic Effects of Tetracycline Analogues (Doxycycline, Minocycline and COL-3) in Acute Myeloid Leukemia HL-60 Cells

    doi: 10.1371/journal.pone.0114457

    Figure Lengend Snippet: TCNAs-induced apoptosis. HL-60 cells were incubated with DOXY (A) and MINO (B) in concentrations of 5, 10, 25 and 50 µg/ml or COL-3 (C) in concentrations of 0.75, 1, and 5 µg/ml until 48 h. Cells incubated in DMSO in a final concentration of 0.2% were used as controls for solvent toxicity and cells incubated in complete medium served as controls. Cells were stained with annexin V and PI for 15 min at room temperature in the dark. Twenty thousand events were acquired and analyzed using flow cytometry. Subpopulations are expressed as percentages of total events. Results are expressed as a mean ± SD of three independent experiments. (D), (E): Examples on flow cytometric analysis of TCNAs-induced apoptosis using annexin V/PI assay. D: DOXY, M: MINO COL: COL-3 and PI: propidium iodide.

    Article Snippet: The other materials were purchased as follows: Z-VAD-FMK (Bachem AG, Bubendorf, Switzerland); etoposide (Bristol-Myers Squibb, Bromma, Sweden); RPMI 1640 medium, Dulbecco’s phosphate-buffer saline (PBS) and fetal bovine serum (FBS) (Invitrogen AB, Stockholm, Sweden); resazurin (R & D System, Stockholm, Sweden); cOmplete mini protease inhibitor cocktail (Roche AB, Stockholm, Sweden); propidium iodide (Sigma Aldrich) and annexin V (BD, Stockholm, Sweden).

    Techniques: Incubation, Concentration Assay, Staining, Flow Cytometry, Cytometry

    The effect of TCNAs on DNA. HL-60 cells were incubated with DOXY or MINO in final concentrations of 25 µg/ml or COL-3 in a final concentration of 5 µg/ml for 10 min and 1, 2 and 4 h. Cells incubated in DMSO in a final concentration of 0.2% were used as controls for solvent toxicity and cells incubated in complete medium served as controls. Cells incubated with etoposide in a final concentration of 6 µg/ml served as positive controls for apoptosis. The TCNAs-induced DNA damage was assessed using SDS-PAGE and immunostaining for the DNA double strand breaks marker γH2AX. Co: control, E: etoposide, D: DOXY, M: MINO and C: COL-3.

    Journal: PLoS ONE

    Article Title: Cytotoxic Effects of Tetracycline Analogues (Doxycycline, Minocycline and COL-3) in Acute Myeloid Leukemia HL-60 Cells

    doi: 10.1371/journal.pone.0114457

    Figure Lengend Snippet: The effect of TCNAs on DNA. HL-60 cells were incubated with DOXY or MINO in final concentrations of 25 µg/ml or COL-3 in a final concentration of 5 µg/ml for 10 min and 1, 2 and 4 h. Cells incubated in DMSO in a final concentration of 0.2% were used as controls for solvent toxicity and cells incubated in complete medium served as controls. Cells incubated with etoposide in a final concentration of 6 µg/ml served as positive controls for apoptosis. The TCNAs-induced DNA damage was assessed using SDS-PAGE and immunostaining for the DNA double strand breaks marker γH2AX. Co: control, E: etoposide, D: DOXY, M: MINO and C: COL-3.

    Article Snippet: The other materials were purchased as follows: Z-VAD-FMK (Bachem AG, Bubendorf, Switzerland); etoposide (Bristol-Myers Squibb, Bromma, Sweden); RPMI 1640 medium, Dulbecco’s phosphate-buffer saline (PBS) and fetal bovine serum (FBS) (Invitrogen AB, Stockholm, Sweden); resazurin (R & D System, Stockholm, Sweden); cOmplete mini protease inhibitor cocktail (Roche AB, Stockholm, Sweden); propidium iodide (Sigma Aldrich) and annexin V (BD, Stockholm, Sweden).

    Techniques: Incubation, Concentration Assay, SDS Page, Immunostaining, Marker

    The loss of mitochondrial membrane potential (Δψm) in TCNAs-induced apoptosis. Mitochondrial membrane potential was assessed using tetramethylrhodamine methyl ester (TMRM) and flow cytometry. (A) HL-60 cells were incubated with DOXY or MINO in final concentrations of 25 and 50 µg/ml or (B) COL-3 in final concentrations of 0.5, 1, 2.5 and 5 µg/ml for 2, 4, 6 and 24 h. Cells incubated in DMSO in a final concentration of 0.2% were used as controls for solvent toxicity and cells incubated in complete medium served as controls. Results are expressed as mean ± SD of three independent experiments. (C), (D): Example on flow cytometric analysis of loss of Δψm using TMRM assay. D: DOXY, M: MINO, COL: COL-3.

    Journal: PLoS ONE

    Article Title: Cytotoxic Effects of Tetracycline Analogues (Doxycycline, Minocycline and COL-3) in Acute Myeloid Leukemia HL-60 Cells

    doi: 10.1371/journal.pone.0114457

    Figure Lengend Snippet: The loss of mitochondrial membrane potential (Δψm) in TCNAs-induced apoptosis. Mitochondrial membrane potential was assessed using tetramethylrhodamine methyl ester (TMRM) and flow cytometry. (A) HL-60 cells were incubated with DOXY or MINO in final concentrations of 25 and 50 µg/ml or (B) COL-3 in final concentrations of 0.5, 1, 2.5 and 5 µg/ml for 2, 4, 6 and 24 h. Cells incubated in DMSO in a final concentration of 0.2% were used as controls for solvent toxicity and cells incubated in complete medium served as controls. Results are expressed as mean ± SD of three independent experiments. (C), (D): Example on flow cytometric analysis of loss of Δψm using TMRM assay. D: DOXY, M: MINO, COL: COL-3.

    Article Snippet: The other materials were purchased as follows: Z-VAD-FMK (Bachem AG, Bubendorf, Switzerland); etoposide (Bristol-Myers Squibb, Bromma, Sweden); RPMI 1640 medium, Dulbecco’s phosphate-buffer saline (PBS) and fetal bovine serum (FBS) (Invitrogen AB, Stockholm, Sweden); resazurin (R & D System, Stockholm, Sweden); cOmplete mini protease inhibitor cocktail (Roche AB, Stockholm, Sweden); propidium iodide (Sigma Aldrich) and annexin V (BD, Stockholm, Sweden).

    Techniques: Flow Cytometry, Cytometry, Incubation, Concentration Assay

    Apocatalase polypeptide proteolysis is induced in late exponential growth phase E. faecalis cells. A , lysates from strain OG1RF cells grown in heme-free medium until early exponential growth phase ( sample A ) or late exponential growth phase ( sample B ) and lysates from cells grown in heme-supplemented medium until late exponential growth phase ( sample C ) were analyzed individually or as mixtures as indicated. The samples were incubated at 37 °C, and the presence of KatA polypeptide in aliquots removed at the indicated time points was determined by immuno-blot. B , proteolysis of KatA in sample A mixed with sample B in the presence of 1 m m PMSF and/or EDTA or Complete EDTA-free protease inhibitor cocktail. The reference, sample A + B without any additions, is indicated by Ctrl .

    Journal: The Journal of Biological Chemistry

    Article Title: In Vitro Assembly of Catalase *

    doi: 10.1074/jbc.M114.596148

    Figure Lengend Snippet: Apocatalase polypeptide proteolysis is induced in late exponential growth phase E. faecalis cells. A , lysates from strain OG1RF cells grown in heme-free medium until early exponential growth phase ( sample A ) or late exponential growth phase ( sample B ) and lysates from cells grown in heme-supplemented medium until late exponential growth phase ( sample C ) were analyzed individually or as mixtures as indicated. The samples were incubated at 37 °C, and the presence of KatA polypeptide in aliquots removed at the indicated time points was determined by immuno-blot. B , proteolysis of KatA in sample A mixed with sample B in the presence of 1 m m PMSF and/or EDTA or Complete EDTA-free protease inhibitor cocktail. The reference, sample A + B without any additions, is indicated by Ctrl .

    Article Snippet: To test protection of KatA by protease inhibitors 1 mm PMSF and/or 1 mm EDTA, or 1× Complete mini EDTA-free protease inhibitor cocktail (Roche Applied Science), were added to lysate from late exponential growth phase cells.

    Techniques: Incubation, Protease Inhibitor