high performance liquid chromatography grade filipin iii  (Millipore)


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    High Performance Liquid Chromatography
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    Structured Review

    Millipore high performance liquid chromatography grade filipin iii
    Cellular uptake of the Arg-nHAP/DZ1 complex in the presence of specific endocytic inhibitors. Cells were pretreated with inhibitors and transfected with the Arg-nHAP/FITC-DZ1 complex. Concentrations of the inhibitors are as follows: 20 mM sodium azide and 50 mM 2-deoxy-D-glucose for one hour; 0.15 μM PAO for 10 minutes; and 1.25 μg/mL of <t>filipin</t> for one hour. The effect of low temperature on cellular uptake was investigated at 4°C. All values are the mean of <t>three</t> measurements and are shown with error bars. Abbreviations: FITC, fluorescein isothiocyanate; Arg-nHAP, arginine-modified nanohydroxyapatite particles; PAO, phenylarsine oxide; DZ1, DNAzyme 1.

    https://www.bioz.com/result/high performance liquid chromatography grade filipin iii/product/Millipore
    Average 99 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    high performance liquid chromatography grade filipin iii - by Bioz Stars, 2020-04
    99/100 stars

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    1) Product Images from "Delivery system for DNAzymes using arginine-modified hydroxyapatite nanoparticles for therapeutic application in a nasopharyngeal carcinoma model"

    Article Title: Delivery system for DNAzymes using arginine-modified hydroxyapatite nanoparticles for therapeutic application in a nasopharyngeal carcinoma model

    Journal: International Journal of Nanomedicine

    doi: 10.2147/IJN.S48321

    Cellular uptake of the Arg-nHAP/DZ1 complex in the presence of specific endocytic inhibitors. Cells were pretreated with inhibitors and transfected with the Arg-nHAP/FITC-DZ1 complex. Concentrations of the inhibitors are as follows: 20 mM sodium azide and 50 mM 2-deoxy-D-glucose for one hour; 0.15 μM PAO for 10 minutes; and 1.25 μg/mL of filipin for one hour. The effect of low temperature on cellular uptake was investigated at 4°C. All values are the mean of three measurements and are shown with error bars. Abbreviations: FITC, fluorescein isothiocyanate; Arg-nHAP, arginine-modified nanohydroxyapatite particles; PAO, phenylarsine oxide; DZ1, DNAzyme 1.
    Figure Legend Snippet: Cellular uptake of the Arg-nHAP/DZ1 complex in the presence of specific endocytic inhibitors. Cells were pretreated with inhibitors and transfected with the Arg-nHAP/FITC-DZ1 complex. Concentrations of the inhibitors are as follows: 20 mM sodium azide and 50 mM 2-deoxy-D-glucose for one hour; 0.15 μM PAO for 10 minutes; and 1.25 μg/mL of filipin for one hour. The effect of low temperature on cellular uptake was investigated at 4°C. All values are the mean of three measurements and are shown with error bars. Abbreviations: FITC, fluorescein isothiocyanate; Arg-nHAP, arginine-modified nanohydroxyapatite particles; PAO, phenylarsine oxide; DZ1, DNAzyme 1.

    Techniques Used: Transfection, Modification

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    Article Snippet: .. The standard (DNJ) for HPLC analysis was obtained from Sigma-Aldrich (St Louis, MO, USA). .. The DNJ content was determined using a validation method established using dilutions of each standard at concentrations ranging from 1.03 to 32.90 ppb injected into the HPLC system (correlation coefficient 0.999).

    Article Title: Development of a Highly Efficient Hybrid Peptide That Increases Immunomodulatory Activity Via the TLR4-Mediated Nuclear Factor-κB Signaling Pathway
    Article Snippet: .. The purity of the peptides was determined to be greater than 95% by reverse-phase high-performance liquid chromatography (Sigma-Aldrich, Singapore) and mass spectrometry (MALDI-TOF MS, Model Autoflex, Bruker Daltonics Inc., Billerica, MA, USA) ( ). .. The peptides were identified by mass spectrometry and dissolved in endotoxin-free water, before being stored at −80 °C.

    Article Title: Active Site Mutations Change the Cleavage Specificity of Neprilysin
    Article Snippet: .. HPLC assays Cleavage of physiological peptides was measured via reverse phase high performance liquid chromatography (HPLC) following incubation of the purified NEP or its mutants with 15 µM insulin B chain (Sigma Aldrich), 24 µM Aß1–40 (Anaspec), or 64 µM leu-ENK (Sigma) in 100 µL of 20 mM MES, pH 6.5, at 37°C. .. HPLC was carried out in a Vydac C4 column using a linear gradient from 0.1% trifluoroacetic acid (TFA) in 95% water, 5% acetonitrile to 0.1% TFA in 50% acetonitrile/water at a flow rate of 1 mL/min.

    Article Title: 3,4-Methylenedioxypyrovalerone prevents while methylone enhances methamphetamine-induced damage to dopamine nerve endings: β-ketoamphetamine modulation of neurotoxicity by the dopamine transporter
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    Article Title: Maternal phenylketonuria syndrome: studies in mice suggest a potential approach to a continuing problem
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    Article Title: Comparisons of Ethanol Extracts of Chinese Propolis (Poplar Type) and Poplar Gums Based on the Antioxidant Activities and Molecular Mechanism
    Article Snippet: .. Materials DPPH, ABTS, Trolox, pinocembrin, gallic acid, quercetin, and the standards used in HPLC analysis were purchased from Sigma (St. Louis, MO, USA). .. Antibodies against HO-1 (lot#:YJ071709CS, catalog#:1922-S, monoclonal), GCLM (catalog#:5529-1, monoclonal), TrxR1 (lot#:YI012101C, catalog#:3928-1, monoclonal), β -tubulin (lot#:YH082302D, catalog#:1879-1, monoclonal), p38 (lot#:YF120305C, catalog#:1544-1, monoclonal), Erk1 Phospho/Erk2 Phospho (lot#:YH072803C, catalog#:2219-1, monoclonal), JNK1 phospho/JNK2 phospho/JNK3 phospho (lot#:YK031401CS, catalog#:3893-S, monoclonal), JNK1 (lot#:YH081203, catalog#:3496-S, monoclonal), AKt1 (lot#:YJ051502DS, catalog#:1081-1, monoclonal), and AKt1 phospho (lot#:YH032209C, catalog#:2118-1, monoclonal) were purchased from Epitomics (Burlingame, CA, USA).

    Article Title: Remodeling of Retinal Fatty Acids in an Animal Model of Diabetes
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    Article Title: Anti-obesity activity, acute toxicity, and chemical constituents of aqueous and ethanol Viola mandshurica extracts
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    Article Title: A Study on Photostability of Amphetamines and Ketamine in Hair Irradiated under Artificial Sunlight
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    Article Title: Sodium butyrate protects the intestinal barrier function in peritonitic mice
    Article Snippet: .. Plasma FD40 was measured by high performance liquid chromatography (HPLC) using fluorescence detection model SIL-20A/SIL-20AC prominence liquid chromatograph (Waters-Millipore) (SHIMADZU Company, Japan). .. The chromatography system for FITC analysis included the following parameters: Autosampler (SIL-20A); RF-10 AXL fluorescence detector set at an excitation and emission wavelength of 494 and 518 nm respectively; Polysep-GFC-P linear size exclusion column (300 × 7.8 mm I.D., Phenomenex, Torrance, CA, USA) was used as the analytical column.

    Flow Cytometry:

    Article Title: Active Site Mutations Change the Cleavage Specificity of Neprilysin
    Article Snippet: HPLC assays Cleavage of physiological peptides was measured via reverse phase high performance liquid chromatography (HPLC) following incubation of the purified NEP or its mutants with 15 µM insulin B chain (Sigma Aldrich), 24 µM Aß1–40 (Anaspec), or 64 µM leu-ENK (Sigma) in 100 µL of 20 mM MES, pH 6.5, at 37°C. .. HPLC was carried out in a Vydac C4 column using a linear gradient from 0.1% trifluoroacetic acid (TFA) in 95% water, 5% acetonitrile to 0.1% TFA in 50% acetonitrile/water at a flow rate of 1 mL/min.

    Permeability:

    Article Title: Sodium butyrate protects the intestinal barrier function in peritonitic mice
    Article Snippet: Paragraph title: Intestinal permeability assay ... Plasma FD40 was measured by high performance liquid chromatography (HPLC) using fluorescence detection model SIL-20A/SIL-20AC prominence liquid chromatograph (Waters-Millipore) (SHIMADZU Company, Japan).

    Filtration:

    Article Title: UHPLC Analysis of Saffron (Crocus sativus L.): Optimization of Separation Using Chemometrics and Detection of Minor Crocetin Esters
    Article Snippet: High performance liquid chromatography-grade methanol and acetonitrile were purchased from Sigma-Aldrich (St. Louis, MO, USA). .. Double deionized water was obtained from a Milli-Q filtration/purification system (Millipore, Bedford, MA, USA).

    Chromatography:

    Article Title: UHPLC Analysis of Saffron (Crocus sativus L.): Optimization of Separation Using Chemometrics and Detection of Minor Crocetin Esters
    Article Snippet: .. High performance liquid chromatography-grade methanol and acetonitrile were purchased from Sigma-Aldrich (St. Louis, MO, USA). .. Double deionized water was obtained from a Milli-Q filtration/purification system (Millipore, Bedford, MA, USA).

    Article Title: Sodium butyrate protects the intestinal barrier function in peritonitic mice
    Article Snippet: Plasma FD40 was measured by high performance liquid chromatography (HPLC) using fluorescence detection model SIL-20A/SIL-20AC prominence liquid chromatograph (Waters-Millipore) (SHIMADZU Company, Japan). .. The chromatography system for FITC analysis included the following parameters: Autosampler (SIL-20A); RF-10 AXL fluorescence detector set at an excitation and emission wavelength of 494 and 518 nm respectively; Polysep-GFC-P linear size exclusion column (300 × 7.8 mm I.D., Phenomenex, Torrance, CA, USA) was used as the analytical column.

    Fluorescence:

    Article Title: Mulberry leaf extract displays antidiabetic activity in db/db mice via Akt and AMP-activated protein kinase phosphorylation
    Article Snippet: Signals were detected at the wavelengths of excitation (254 nm) and emission (322 nm) using a fluorescence detector. .. The standard (DNJ) for HPLC analysis was obtained from Sigma-Aldrich (St Louis, MO, USA).

    Article Title: Sodium butyrate protects the intestinal barrier function in peritonitic mice
    Article Snippet: .. Plasma FD40 was measured by high performance liquid chromatography (HPLC) using fluorescence detection model SIL-20A/SIL-20AC prominence liquid chromatograph (Waters-Millipore) (SHIMADZU Company, Japan). .. The chromatography system for FITC analysis included the following parameters: Autosampler (SIL-20A); RF-10 AXL fluorescence detector set at an excitation and emission wavelength of 494 and 518 nm respectively; Polysep-GFC-P linear size exclusion column (300 × 7.8 mm I.D., Phenomenex, Torrance, CA, USA) was used as the analytical column.

    Synthesized:

    Article Title: Development of a Highly Efficient Hybrid Peptide That Increases Immunomodulatory Activity Via the TLR4-Mediated Nuclear Factor-κB Signaling Pathway
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    Multiple Displacement Amplification:

    Article Title: A Study on Photostability of Amphetamines and Ketamine in Hair Irradiated under Artificial Sunlight
    Article Snippet: The internal standards (IS) MA-D5, AMF-D5, MDMA-D5, MDA-D5, and KET-D3 were methanol solutions also from LGC Promochem at 1.0 mg/mL. .. High-performance liquid chromatography water was prepared using a Milli-Q Plus (Millipore, Molsheim, France) system.

    Bicinchoninic Acid Protein Assay:

    Article Title: 3,4-Methylenedioxypyrovalerone prevents while methylone enhances methamphetamine-induced damage to dopamine nerve endings: β-ketoamphetamine modulation of neurotoxicity by the dopamine transporter
    Article Snippet: Methylone hydrochloride, MDPV hydrochloride, (±) mephedrone hydrochloride, and (±) MDMA hydrochloride were obtained from the NIDA Research Resources Drug Supply Program. (+) methamphetamine hydrochloride, D-amphetamine sulfate, MPTP hydrochloride, DA, polyclonal antibodies against glyceraldehyde 3-phosphate dehydrogenase (GAPDH), and all buffers and HPLC reagents were purchased from Sigma-Aldrich (St. Louis, MO, USA). .. Bicinchoninic acid protein assay kits were obtained from Pierce (Rockford, IL, USA).

    Purification:

    Article Title: Development of a Highly Efficient Hybrid Peptide That Increases Immunomodulatory Activity Via the TLR4-Mediated Nuclear Factor-κB Signaling Pathway
    Article Snippet: Peptide Synthesis The hybrid peptides LTAa , LTAb , and LTAc and their parental peptides LL-37 and Tα1 were synthesized and purified by KangLong Biochemistry (Jiangsu, China) using solid-phase methods. .. The purity of the peptides was determined to be greater than 95% by reverse-phase high-performance liquid chromatography (Sigma-Aldrich, Singapore) and mass spectrometry (MALDI-TOF MS, Model Autoflex, Bruker Daltonics Inc., Billerica, MA, USA) ( ).

    Article Title: Active Site Mutations Change the Cleavage Specificity of Neprilysin
    Article Snippet: .. HPLC assays Cleavage of physiological peptides was measured via reverse phase high performance liquid chromatography (HPLC) following incubation of the purified NEP or its mutants with 15 µM insulin B chain (Sigma Aldrich), 24 µM Aß1–40 (Anaspec), or 64 µM leu-ENK (Sigma) in 100 µL of 20 mM MES, pH 6.5, at 37°C. .. HPLC was carried out in a Vydac C4 column using a linear gradient from 0.1% trifluoroacetic acid (TFA) in 95% water, 5% acetonitrile to 0.1% TFA in 50% acetonitrile/water at a flow rate of 1 mL/min.

    Produced:

    Article Title: 3,4-Methylenedioxypyrovalerone prevents while methylone enhances methamphetamine-induced damage to dopamine nerve endings: β-ketoamphetamine modulation of neurotoxicity by the dopamine transporter
    Article Snippet: Methylone hydrochloride, MDPV hydrochloride, (±) mephedrone hydrochloride, and (±) MDMA hydrochloride were obtained from the NIDA Research Resources Drug Supply Program. (+) methamphetamine hydrochloride, D-amphetamine sulfate, MPTP hydrochloride, DA, polyclonal antibodies against glyceraldehyde 3-phosphate dehydrogenase (GAPDH), and all buffers and HPLC reagents were purchased from Sigma-Aldrich (St. Louis, MO, USA). .. Polyclonal antibodies against rat tyrosine hydroxylase (TH) were produced as described previously ( ).

    Article Title: UHPLC Analysis of Saffron (Crocus sativus L.): Optimization of Separation Using Chemometrics and Detection of Minor Crocetin Esters
    Article Snippet: Samples, Chemicals and Solvents Saffron samples in stigmas produced in L’Aquila (Abruzzo, Italy), Morocco (Taliouine) and Iran (Khorasan province) in 2015 were analyzed. .. High performance liquid chromatography-grade methanol and acetonitrile were purchased from Sigma-Aldrich (St. Louis, MO, USA).

    Incubation:

    Article Title: Active Site Mutations Change the Cleavage Specificity of Neprilysin
    Article Snippet: .. HPLC assays Cleavage of physiological peptides was measured via reverse phase high performance liquid chromatography (HPLC) following incubation of the purified NEP or its mutants with 15 µM insulin B chain (Sigma Aldrich), 24 µM Aß1–40 (Anaspec), or 64 µM leu-ENK (Sigma) in 100 µL of 20 mM MES, pH 6.5, at 37°C. .. HPLC was carried out in a Vydac C4 column using a linear gradient from 0.1% trifluoroacetic acid (TFA) in 95% water, 5% acetonitrile to 0.1% TFA in 50% acetonitrile/water at a flow rate of 1 mL/min.

    Mass Spectrometry:

    Article Title: Development of a Highly Efficient Hybrid Peptide That Increases Immunomodulatory Activity Via the TLR4-Mediated Nuclear Factor-κB Signaling Pathway
    Article Snippet: .. The purity of the peptides was determined to be greater than 95% by reverse-phase high-performance liquid chromatography (Sigma-Aldrich, Singapore) and mass spectrometry (MALDI-TOF MS, Model Autoflex, Bruker Daltonics Inc., Billerica, MA, USA) ( ). .. The peptides were identified by mass spectrometry and dissolved in endotoxin-free water, before being stored at −80 °C.

    Sampling:

    Article Title: Sodium butyrate protects the intestinal barrier function in peritonitic mice
    Article Snippet: Animals were gavaged with 60 mg/kg FD40 4 h prior to blood sampling from the orbital sinus. .. Plasma FD40 was measured by high performance liquid chromatography (HPLC) using fluorescence detection model SIL-20A/SIL-20AC prominence liquid chromatograph (Waters-Millipore) (SHIMADZU Company, Japan).

    Molecular Weight:

    Article Title: Sodium butyrate protects the intestinal barrier function in peritonitic mice
    Article Snippet: Fluorescein isothiocyanate-dextran, molecular weight 40 kD, (FD40), (Sigma-Aldrich, USA), was used as a tracer of intestinal permeability. .. Plasma FD40 was measured by high performance liquid chromatography (HPLC) using fluorescence detection model SIL-20A/SIL-20AC prominence liquid chromatograph (Waters-Millipore) (SHIMADZU Company, Japan).

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    Millipore filipin
    StarD5 deletion lowers PM cholesterol in macrophages and ABCA1-dependent cholesterol efflux. A: Primary peritoneal macrophages from WT and StarD5 −/− mice were imaged following staining with <t>filipin</t> and BODIPY 493-503, as indicated, as well as by differential interference contrast for those stained with BODIPY 493-503. B: Total, free, esterified, and PM cholesterol were quantified as described in the Materials and Methods (n = 4). C: Accessible PM cholesterol was quantified as described in the Materials and Methods (n = 3, * P ≤ 0.05 for WT versus StarD5 −/− ). D: Primary peritoneal macrophages from WT and StarD5 −/− mice were used in cholesterol efflux assays as described in the Materials and Methods in the absence or presence of cAMP or cyclosporine A. Cholesterol efflux was calculated as the percentage of total cell 3 H-cholesterol content, which was about 5% of the total cholesterol (total effluxed 3 H-cholesterol + cell-associated 3 H-cholesterol) and represented as a percentage of the effluxed cholesterol in WT macrophages in the presence of cAMP. Primary peritoneal macrophages from StarD5 −/− mice infected with Ad-StarD5 (MOI of 200) were also used. The values represent the average ± SD (n = 4, * P
    Filipin, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 83 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/filipin/product/Millipore
    Average 99 stars, based on 83 article reviews
    Price from $9.99 to $1999.99
    filipin - by Bioz Stars, 2020-04
    99/100 stars
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    StarD5 deletion lowers PM cholesterol in macrophages and ABCA1-dependent cholesterol efflux. A: Primary peritoneal macrophages from WT and StarD5 −/− mice were imaged following staining with filipin and BODIPY 493-503, as indicated, as well as by differential interference contrast for those stained with BODIPY 493-503. B: Total, free, esterified, and PM cholesterol were quantified as described in the Materials and Methods (n = 4). C: Accessible PM cholesterol was quantified as described in the Materials and Methods (n = 3, * P ≤ 0.05 for WT versus StarD5 −/− ). D: Primary peritoneal macrophages from WT and StarD5 −/− mice were used in cholesterol efflux assays as described in the Materials and Methods in the absence or presence of cAMP or cyclosporine A. Cholesterol efflux was calculated as the percentage of total cell 3 H-cholesterol content, which was about 5% of the total cholesterol (total effluxed 3 H-cholesterol + cell-associated 3 H-cholesterol) and represented as a percentage of the effluxed cholesterol in WT macrophages in the presence of cAMP. Primary peritoneal macrophages from StarD5 −/− mice infected with Ad-StarD5 (MOI of 200) were also used. The values represent the average ± SD (n = 4, * P

    Journal: Journal of Lipid Research

    Article Title: StarD5: an ER stress protein regulates plasma membrane and intracellular cholesterol homeostasis [S]

    doi: 10.1194/jlr.M091967

    Figure Lengend Snippet: StarD5 deletion lowers PM cholesterol in macrophages and ABCA1-dependent cholesterol efflux. A: Primary peritoneal macrophages from WT and StarD5 −/− mice were imaged following staining with filipin and BODIPY 493-503, as indicated, as well as by differential interference contrast for those stained with BODIPY 493-503. B: Total, free, esterified, and PM cholesterol were quantified as described in the Materials and Methods (n = 4). C: Accessible PM cholesterol was quantified as described in the Materials and Methods (n = 3, * P ≤ 0.05 for WT versus StarD5 −/− ). D: Primary peritoneal macrophages from WT and StarD5 −/− mice were used in cholesterol efflux assays as described in the Materials and Methods in the absence or presence of cAMP or cyclosporine A. Cholesterol efflux was calculated as the percentage of total cell 3 H-cholesterol content, which was about 5% of the total cholesterol (total effluxed 3 H-cholesterol + cell-associated 3 H-cholesterol) and represented as a percentage of the effluxed cholesterol in WT macrophages in the presence of cAMP. Primary peritoneal macrophages from StarD5 −/− mice infected with Ad-StarD5 (MOI of 200) were also used. The values represent the average ± SD (n = 4, * P

    Article Snippet: Filipin was from Cayman Biochemicals; cholesterol oxidase was from Calbiochem, BODIPY 493/503 was from Fisher Scientific; cAMP, cyclosporin A, and other general chemicals were from Sigma.

    Techniques: Mouse Assay, Staining, Infection

    Disruption of lipid rafts inhibits the antiproliferative effects of GPR55 activation. ( A ) Mz-ChA-1 cells were pretreated with lipid raft disrupters 0.1 mM β–methylcyclodextrin, or 1 μg/mL filipin III for 1 hr prior to the addition of O-1602 (10 −5 M). Cell viability was determined by MTS assays. Data are expressed as average ± SEM (*p

    Journal: Laboratory investigation; a journal of technical methods and pathology

    Article Title: Anandamide exerts its antiproliferative actions on cholangiocarcinoma by activation of the GPR55 receptor

    doi: 10.1038/labinvest.2011.62

    Figure Lengend Snippet: Disruption of lipid rafts inhibits the antiproliferative effects of GPR55 activation. ( A ) Mz-ChA-1 cells were pretreated with lipid raft disrupters 0.1 mM β–methylcyclodextrin, or 1 μg/mL filipin III for 1 hr prior to the addition of O-1602 (10 −5 M). Cell viability was determined by MTS assays. Data are expressed as average ± SEM (*p

    Article Snippet: The inhibitors utilized were the lipid raft disruptors, methyl-β-cyclodextrin (0.1 mM), , and filipin III (1 mg/mL) ( , ), and the JNK inhibitor (10−7 M; SP600125; EMD Chemicals, Gibbstown, NJ) ( ).

    Techniques: Activation Assay

    Overall structures of CYP105P1-filipin I complex ( A ) and unliganded CYP105D6 ( B ), illustrated by ribbon representation. Heme and ligands are shown as stick models. The BC and FG loop regions are shown in dark gray. a.a. , amino acids.

    Journal: The Journal of Biological Chemistry

    Article Title: Regio- and Stereospecificity of Filipin Hydroxylation Sites Revealed by Crystal Structures of Cytochrome P450 105P1 and 105D6 from Streptomyces avermitilis *

    doi: 10.1074/jbc.M109.092460

    Figure Lengend Snippet: Overall structures of CYP105P1-filipin I complex ( A ) and unliganded CYP105D6 ( B ), illustrated by ribbon representation. Heme and ligands are shown as stick models. The BC and FG loop regions are shown in dark gray. a.a. , amino acids.

    Article Snippet: Before crystallization, filipin I was dissolved in dimethyl sulfoxide, mixed with a CYP105P1 sample, and concentrated using an Ultrafree Centrifugal Filter Device (Millipore, Billerica, MA).

    Techniques:

    Spectral changes of CYP105P1 (ferric resting state) upon the addition of increasing concentrations of filipin I ( A ), its difference spectra ( B ), and the titration curve calculated using the values of absorption differences at 387 and 422 nm ( C ).

    Journal: The Journal of Biological Chemistry

    Article Title: Regio- and Stereospecificity of Filipin Hydroxylation Sites Revealed by Crystal Structures of Cytochrome P450 105P1 and 105D6 from Streptomyces avermitilis *

    doi: 10.1074/jbc.M109.092460

    Figure Lengend Snippet: Spectral changes of CYP105P1 (ferric resting state) upon the addition of increasing concentrations of filipin I ( A ), its difference spectra ( B ), and the titration curve calculated using the values of absorption differences at 387 and 422 nm ( C ).

    Article Snippet: Before crystallization, filipin I was dissolved in dimethyl sulfoxide, mixed with a CYP105P1 sample, and concentrated using an Ultrafree Centrifugal Filter Device (Millipore, Billerica, MA).

    Techniques: Titration

    Interactions between CYP105P1 and filipin I. A , shown is an F obs − F calc omit electron density map of the filipin I molecule contoured at 4.0 σ. B , hydrophobic interactions are observed at both sides of the 28-membered ring. Labels for atoms of filipin I are underlined. C , a stereographic figure shows interactions with the BC loop, FG helices, and I helix. The water molecules mediate hydrophilic interactions with the polyol group of filipin I. The extensive hydrogen-bonding network and residues involved in it are shown as dotted lines and stick models, respectively. The distance between the C26 atom of filipin I and the heme iron is 5.0 Å, and the pro-S hydrogen side is directed toward the heme.

    Journal: The Journal of Biological Chemistry

    Article Title: Regio- and Stereospecificity of Filipin Hydroxylation Sites Revealed by Crystal Structures of Cytochrome P450 105P1 and 105D6 from Streptomyces avermitilis *

    doi: 10.1074/jbc.M109.092460

    Figure Lengend Snippet: Interactions between CYP105P1 and filipin I. A , shown is an F obs − F calc omit electron density map of the filipin I molecule contoured at 4.0 σ. B , hydrophobic interactions are observed at both sides of the 28-membered ring. Labels for atoms of filipin I are underlined. C , a stereographic figure shows interactions with the BC loop, FG helices, and I helix. The water molecules mediate hydrophilic interactions with the polyol group of filipin I. The extensive hydrogen-bonding network and residues involved in it are shown as dotted lines and stick models, respectively. The distance between the C26 atom of filipin I and the heme iron is 5.0 Å, and the pro-S hydrogen side is directed toward the heme.

    Article Snippet: Before crystallization, filipin I was dissolved in dimethyl sulfoxide, mixed with a CYP105P1 sample, and concentrated using an Ultrafree Centrifugal Filter Device (Millipore, Billerica, MA).

    Techniques:

    Active site structures of CYP105P1-filipin I ( green ) superimposed with CYP105D6 ( A , cyan ), P450cam-camphor-O 2 ( B , yellow ), and P450 EryK-erythromycin D ( C , magenta ). Water molecules and the hydroxylation target positions of substrates (C26 of filipin I, C5 of camphor, and C12 of erythromycin D) are shown as spheres . A water molecule is positioned 2.8 Å from the heme iron in the CYP105D6 structure ( A ).

    Journal: The Journal of Biological Chemistry

    Article Title: Regio- and Stereospecificity of Filipin Hydroxylation Sites Revealed by Crystal Structures of Cytochrome P450 105P1 and 105D6 from Streptomyces avermitilis *

    doi: 10.1074/jbc.M109.092460

    Figure Lengend Snippet: Active site structures of CYP105P1-filipin I ( green ) superimposed with CYP105D6 ( A , cyan ), P450cam-camphor-O 2 ( B , yellow ), and P450 EryK-erythromycin D ( C , magenta ). Water molecules and the hydroxylation target positions of substrates (C26 of filipin I, C5 of camphor, and C12 of erythromycin D) are shown as spheres . A water molecule is positioned 2.8 Å from the heme iron in the CYP105D6 structure ( A ).

    Article Snippet: Before crystallization, filipin I was dissolved in dimethyl sulfoxide, mixed with a CYP105P1 sample, and concentrated using an Ultrafree Centrifugal Filter Device (Millipore, Billerica, MA).

    Techniques:

    Closing motion of CYP105P1. Shown is a stereographic superimposition of the filipin I complex structure with the ligand-free wild-type ( A ) and H72A mutant ( B ) structures. BC loop and FG helices are colored magenta and green in ligand-free and complex structures, respectively. The filipin I molecule is shown as yellow sticks . In the ligand-free wild-type structure ( A ), the side chain of His-72 is ligated to the heme iron as the sixth ligand, and the BC loop sinks into the heme to completely cover the distal pocket.

    Journal: The Journal of Biological Chemistry

    Article Title: Regio- and Stereospecificity of Filipin Hydroxylation Sites Revealed by Crystal Structures of Cytochrome P450 105P1 and 105D6 from Streptomyces avermitilis *

    doi: 10.1074/jbc.M109.092460

    Figure Lengend Snippet: Closing motion of CYP105P1. Shown is a stereographic superimposition of the filipin I complex structure with the ligand-free wild-type ( A ) and H72A mutant ( B ) structures. BC loop and FG helices are colored magenta and green in ligand-free and complex structures, respectively. The filipin I molecule is shown as yellow sticks . In the ligand-free wild-type structure ( A ), the side chain of His-72 is ligated to the heme iron as the sixth ligand, and the BC loop sinks into the heme to completely cover the distal pocket.

    Article Snippet: Before crystallization, filipin I was dissolved in dimethyl sulfoxide, mixed with a CYP105P1 sample, and concentrated using an Ultrafree Centrifugal Filter Device (Millipore, Billerica, MA).

    Techniques: Mutagenesis

    Spectral changes of CYP105D6 (ferric resting state) upon the addition of increasing concentrations of filipin I ( A ), its difference spectra ( B ), and the titration curve calculated using the values of absorption differences at 387 and 420 nm ( C ). A nonlinear fitting with a quadratic equation was applied to the titration curve.

    Journal: The Journal of Biological Chemistry

    Article Title: Regio- and Stereospecificity of Filipin Hydroxylation Sites Revealed by Crystal Structures of Cytochrome P450 105P1 and 105D6 from Streptomyces avermitilis *

    doi: 10.1074/jbc.M109.092460

    Figure Lengend Snippet: Spectral changes of CYP105D6 (ferric resting state) upon the addition of increasing concentrations of filipin I ( A ), its difference spectra ( B ), and the titration curve calculated using the values of absorption differences at 387 and 420 nm ( C ). A nonlinear fitting with a quadratic equation was applied to the titration curve.

    Article Snippet: Before crystallization, filipin I was dissolved in dimethyl sulfoxide, mixed with a CYP105P1 sample, and concentrated using an Ultrafree Centrifugal Filter Device (Millipore, Billerica, MA).

    Techniques: Titration

    Internalization of the hMCRs in HEK293 cells. Cells were incubated with [ 125 I]-NDP- α -MSH and caveolae inhibitors, filipin (▼) and nystatin (▼) compared with control (|) at 37 °C for the indicated times. Acid-resistant and acid-sensitive binding were determined. Percentage internalization (acid-resistant binding) was expressed by measuring total binding at each time-point.

    Journal: Chemical biology & drug design

    Article Title: Cell Signaling and Trafficking of Human Melanocortin Receptors in Real Time Using Two-photon Fluorescence and Confocal Laser Microscopy: Differentiation of Agonists and Antagonists

    doi: 10.1111/j.1747-0285.2006.00432.x

    Figure Lengend Snippet: Internalization of the hMCRs in HEK293 cells. Cells were incubated with [ 125 I]-NDP- α -MSH and caveolae inhibitors, filipin (▼) and nystatin (▼) compared with control (|) at 37 °C for the indicated times. Acid-resistant and acid-sensitive binding were determined. Percentage internalization (acid-resistant binding) was expressed by measuring total binding at each time-point.

    Article Snippet: H-89 and Filipin were from Calbiochem (San Diego, CA, USA).

    Techniques: Incubation, Binding Assay