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  • 99
    Thermo Fisher fatty acid free bovine serum albumin bsa
    Fatty Acid Free Bovine Serum Albumin Bsa, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 42 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore bsa free fatty acids
    Bsa Free Fatty Acids, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore fatty acid free bovine serum album bsa
    Modulation of S1P homeostasis of K562 cells by sodium butyrate-induced differentiation into erythroblast-like cells. K562 cells were treated with 2 mM sodium butyrate (NaB); 72 hours later, we investigated the modulation of S1P homeostasis. (A) The mRNA levels of GYPA and ALAS were determined using real-time PCR. GAPDH was utilized as an internal control (n = 8/group). (B, C) The expression of CD235a was investigated with a flow cytometer (n = 3/group). (D) C 17 S1P formation assay. NaB-treated or vehicle-treated K562 cells were treated with 10 μM of C 17 sphingosine for 20 minutes. Then, we replaced the supernatant with <t>PBS</t> containing 0.5% <t>BSA</t> and incubated the cells at 37°C for another 20 minutes. Then, the supernatants and cells were collected and used for the C 17 S1P measurements (n = 6/group). (E) The expression of key enzymes in S1P metabolism was determined using real-time PCR. GAPDH was utilized as an internal control (n = 8/group). (F) SK activity assay. The SK activity assay was performed using NaB-treated or vehicle-treated K562 cells (n = 6/group). (G) Reverse transcription PCR was performed using cDNAs prepared from NaB-treated K562 cells (K), HepG2 cells (He), and HUVECs (Hu). (H) The expression of possible S1P transporters was determined using real-time PCR. GAPDH was utilized as an internal control (n = 4/group). (I) Real-time PCR of Band3. GAPDH was utilized as an internal control (n = 8/group). (J) Western blot of Band3 with membranous protein. The whole cell lysate of RBCs (2 μg) was placed as a positive control. Pan-cadherin was utilized as an internal control (n = 3/group).
    Fatty Acid Free Bovine Serum Album Bsa, supplied by Millipore, used in various techniques. Bioz Stars score: 90/100, based on 414 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    96
    Roche fatty acid free bsa
    p21-dependent inhibition of <t>LPA-induced</t> cell proliferation by TGFβ. A . TGFβ inhibits LPA-afforded cell proliferation. MDA-MB-231 and Caov-3 cells in 6-well plates were incubated with LPA (10 µM) or vehicle <t>(BSA)</t> in the presence
    Fatty Acid Free Bsa, supplied by Roche, used in various techniques. Bioz Stars score: 96/100, based on 234 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Boehringer Mannheim fatty acid free bsa
    p21-dependent inhibition of <t>LPA-induced</t> cell proliferation by TGFβ. A . TGFβ inhibits LPA-afforded cell proliferation. MDA-MB-231 and Caov-3 cells in 6-well plates were incubated with LPA (10 µM) or vehicle <t>(BSA)</t> in the presence
    Fatty Acid Free Bsa, supplied by Boehringer Mannheim, used in various techniques. Bioz Stars score: 86/100, based on 77 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Equitech-Bio fatty acid free bsa
    Effect of diabetes on cholesterol efflux to <t>apoA1</t> or HDL and regulation of cholesterol transporters by Ager in primary murine BMDMs. Primary BMDMs were retrieved from nondiabetic and diabetic WT (2 months of hyperglycemia) and Ager −/− mice and were cultured in concentrations of glucose consistent with the glycemic state of the mice from which they were retrieved. BMDMs were labeled with 3 H-cholesterol and treated with acetylated LDL and fatty acid–free <t>BSA</t> (1%) for 24 h. A and B : To mediate cholesterol efflux, cells were treated with apoA1 (5 μg/mL) ( A ) or HDL (100 μg/mL) ( B ) for 6 h, and supernatant was collected. Percent cholesterol efflux is the radioactivity in the supernatant divided by the sum of the radioactivity measured in the supernatant and the cell lysates from n ≥ 5 mice/group. C and D : BMDMs were retrieved from WT or Ager −/− mice and subjected to quantitative real-time PCR for detection of Abca1 mRNA transcript ( C ) and ABCA1 protein ( D ) analysis ( n = 3 mice/group). E and F : BMDMs were prepared as in C and D and assessed for levels of Abcg1 mRNA transcripts ( E ) and ABCG1 protein ( F ) ( n = 3 mice/group). Error bars represent mean ± SEM. hr, hour; NS, not statistically significant.
    Fatty Acid Free Bsa, supplied by Equitech-Bio, used in various techniques. Bioz Stars score: 94/100, based on 57 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    FUJIFILM fatty acid free bsa
    C-terminus of ATP11C is required for inhibition of flippase activities by PKC activation. a – c Ba/F3 cells stably expressing HA-tagged ATP11A, ATP11C, and ATP11AAC chimeric proteins were treated with vehicle alone ( a ), 400 nM PMA ( b ), or 1 μM A23187 in the presence of CaCl 2 ( c ) for 15 min. The cells were fixed and immunostained with anti-HA and anti-ATP1A1 antibodies, followed by incubation with Cy3-conjugated anti-rat and Alexa Fluor 488-conjugated anti-rabbit secondary antibodies. Scale bars, 5 μm. d – f Ba/F3 cells were treated with vehicle alone (white bars), 400 nM PMA (black bars), or 400 nM PMA and 2 μM of BIM-1 (gray bars) for 15 min. The cells were then washed with flippase assay buffer and incubated at 15 °C for 5 min with <t>NBD-PS</t> ( d ), or 15 min with NBD-PE ( e ) or NBD-PC ( f ). After extraction with fatty acid-free <t>BSA,</t> the residual fluorescence intensity associated with the cells was determined by flow cytometry. Fold increase of NBD-lipids uptake is shown relative to that in Mock-treated parental Ba/F3 cells (−). Graphs display averages from three independent experiments ± SD. ** p
    Fatty Acid Free Bsa, supplied by FUJIFILM, used in various techniques. Bioz Stars score: 93/100, based on 65 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Carl Roth GmbH fatty acid free bsa
    C-terminus of ATP11C is required for inhibition of flippase activities by PKC activation. a – c Ba/F3 cells stably expressing HA-tagged ATP11A, ATP11C, and ATP11AAC chimeric proteins were treated with vehicle alone ( a ), 400 nM PMA ( b ), or 1 μM A23187 in the presence of CaCl 2 ( c ) for 15 min. The cells were fixed and immunostained with anti-HA and anti-ATP1A1 antibodies, followed by incubation with Cy3-conjugated anti-rat and Alexa Fluor 488-conjugated anti-rabbit secondary antibodies. Scale bars, 5 μm. d – f Ba/F3 cells were treated with vehicle alone (white bars), 400 nM PMA (black bars), or 400 nM PMA and 2 μM of BIM-1 (gray bars) for 15 min. The cells were then washed with flippase assay buffer and incubated at 15 °C for 5 min with <t>NBD-PS</t> ( d ), or 15 min with NBD-PE ( e ) or NBD-PC ( f ). After extraction with fatty acid-free <t>BSA,</t> the residual fluorescence intensity associated with the cells was determined by flow cytometry. Fold increase of NBD-lipids uptake is shown relative to that in Mock-treated parental Ba/F3 cells (−). Graphs display averages from three independent experiments ± SD. ** p
    Fatty Acid Free Bsa, supplied by Carl Roth GmbH, used in various techniques. Bioz Stars score: 92/100, based on 21 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Jackson Immuno fatty acid free bsa
    C-terminus of ATP11C is required for inhibition of flippase activities by PKC activation. a – c Ba/F3 cells stably expressing HA-tagged ATP11A, ATP11C, and ATP11AAC chimeric proteins were treated with vehicle alone ( a ), 400 nM PMA ( b ), or 1 μM A23187 in the presence of CaCl 2 ( c ) for 15 min. The cells were fixed and immunostained with anti-HA and anti-ATP1A1 antibodies, followed by incubation with Cy3-conjugated anti-rat and Alexa Fluor 488-conjugated anti-rabbit secondary antibodies. Scale bars, 5 μm. d – f Ba/F3 cells were treated with vehicle alone (white bars), 400 nM PMA (black bars), or 400 nM PMA and 2 μM of BIM-1 (gray bars) for 15 min. The cells were then washed with flippase assay buffer and incubated at 15 °C for 5 min with <t>NBD-PS</t> ( d ), or 15 min with NBD-PE ( e ) or NBD-PC ( f ). After extraction with fatty acid-free <t>BSA,</t> the residual fluorescence intensity associated with the cells was determined by flow cytometry. Fold increase of NBD-lipids uptake is shown relative to that in Mock-treated parental Ba/F3 cells (−). Graphs display averages from three independent experiments ± SD. ** p
    Fatty Acid Free Bsa, supplied by Jackson Immuno, used in various techniques. Bioz Stars score: 95/100, based on 19 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Valiant fatty acid free bsa
    Analyses of flippase activity in different human cancer cell types A. Flippase activity assay: Indicated cell types were incubated with <t>NBD-PS</t> for indicated time periods and subjected to <t>BSA</t> extraction and sodium dithionite treatment. % nonextractable NBD-PS (after BSA extraction and sodium dithionite treatment) represents internalized NBD-PS, indicative of flippase activity (Schwann ◊, U87ΔEGFR-Luc □, H1299 Δ, MDA-MB-231 ○, MDA-MB-231-Luc-D3H2LN ◆, Gli36 ▲, U373 ●) B. NBD PS incorporation rate. C. NBD PS incorporation at 15 minutes time point.
    Fatty Acid Free Bsa, supplied by Valiant, used in various techniques. Bioz Stars score: 92/100, based on 65 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    PAA Laboratories fatty acid free bsa
    Analyses of flippase activity in different human cancer cell types A. Flippase activity assay: Indicated cell types were incubated with <t>NBD-PS</t> for indicated time periods and subjected to <t>BSA</t> extraction and sodium dithionite treatment. % nonextractable NBD-PS (after BSA extraction and sodium dithionite treatment) represents internalized NBD-PS, indicative of flippase activity (Schwann ◊, U87ΔEGFR-Luc □, H1299 Δ, MDA-MB-231 ○, MDA-MB-231-Luc-D3H2LN ◆, Gli36 ▲, U373 ●) B. NBD PS incorporation rate. C. NBD PS incorporation at 15 minutes time point.
    Fatty Acid Free Bsa, supplied by PAA Laboratories, used in various techniques. Bioz Stars score: 91/100, based on 43 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Akron Biotech fatty acid free bsa
    Analyses of flippase activity in different human cancer cell types A. Flippase activity assay: Indicated cell types were incubated with <t>NBD-PS</t> for indicated time periods and subjected to <t>BSA</t> extraction and sodium dithionite treatment. % nonextractable NBD-PS (after BSA extraction and sodium dithionite treatment) represents internalized NBD-PS, indicative of flippase activity (Schwann ◊, U87ΔEGFR-Luc □, H1299 Δ, MDA-MB-231 ○, MDA-MB-231-Luc-D3H2LN ◆, Gli36 ▲, U373 ●) B. NBD PS incorporation rate. C. NBD PS incorporation at 15 minutes time point.
    Fatty Acid Free Bsa, supplied by Akron Biotech, used in various techniques. Bioz Stars score: 93/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Amresco fatty acid free bsa
    Analyses of flippase activity in different human cancer cell types A. Flippase activity assay: Indicated cell types were incubated with <t>NBD-PS</t> for indicated time periods and subjected to <t>BSA</t> extraction and sodium dithionite treatment. % nonextractable NBD-PS (after BSA extraction and sodium dithionite treatment) represents internalized NBD-PS, indicative of flippase activity (Schwann ◊, U87ΔEGFR-Luc □, H1299 Δ, MDA-MB-231 ○, MDA-MB-231-Luc-D3H2LN ◆, Gli36 ▲, U373 ●) B. NBD PS incorporation rate. C. NBD PS incorporation at 15 minutes time point.
    Fatty Acid Free Bsa, supplied by Amresco, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GE Healthcare fatty acid free bsa
    The effect of VASP on lipid metabolism in <t>AML12</t> cells. A : AML12 cells were transduced with VASP (VASP-OE) or control (ctl) (empty) vector. B : RT-PCR analysis of lipid metabolism–related genes in AML12 cells. Expression of Gapdh as shown by the ratio of CT value ( n = 3). C : Rate of [1- 14 C]palmitate incorporation into acid-soluble metabolites ( n = 4). D : Oleic acid (either 0.1 or 0.4 mmol/L)-induced accumulation of TG in AML12 cells. <t>BSA</t> (1.3 mmol/L) was used as a control ( n = 3). * P
    Fatty Acid Free Bsa, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 88/100, based on 89 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Fisher Scientific fatty acid free bsa
    The effect of VASP on lipid metabolism in <t>AML12</t> cells. A : AML12 cells were transduced with VASP (VASP-OE) or control (ctl) (empty) vector. B : RT-PCR analysis of lipid metabolism–related genes in AML12 cells. Expression of Gapdh as shown by the ratio of CT value ( n = 3). C : Rate of [1- 14 C]palmitate incorporation into acid-soluble metabolites ( n = 4). D : Oleic acid (either 0.1 or 0.4 mmol/L)-induced accumulation of TG in AML12 cells. <t>BSA</t> (1.3 mmol/L) was used as a control ( n = 3). * P
    Fatty Acid Free Bsa, supplied by Fisher Scientific, used in various techniques. Bioz Stars score: 92/100, based on 21 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    SeraCare fatty acid free bsa
    The effect of VASP on lipid metabolism in <t>AML12</t> cells. A : AML12 cells were transduced with VASP (VASP-OE) or control (ctl) (empty) vector. B : RT-PCR analysis of lipid metabolism–related genes in AML12 cells. Expression of Gapdh as shown by the ratio of CT value ( n = 3). C : Rate of [1- 14 C]palmitate incorporation into acid-soluble metabolites ( n = 4). D : Oleic acid (either 0.1 or 0.4 mmol/L)-induced accumulation of TG in AML12 cells. <t>BSA</t> (1.3 mmol/L) was used as a control ( n = 3). * P
    Fatty Acid Free Bsa, supplied by SeraCare, used in various techniques. Bioz Stars score: 92/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Applygen Technologies fatty acid free bsa
    The effect of VASP on lipid metabolism in <t>AML12</t> cells. A : AML12 cells were transduced with VASP (VASP-OE) or control (ctl) (empty) vector. B : RT-PCR analysis of lipid metabolism–related genes in AML12 cells. Expression of Gapdh as shown by the ratio of CT value ( n = 3). C : Rate of [1- 14 C]palmitate incorporation into acid-soluble metabolites ( n = 4). D : Oleic acid (either 0.1 or 0.4 mmol/L)-induced accumulation of TG in AML12 cells. <t>BSA</t> (1.3 mmol/L) was used as a control ( n = 3). * P
    Fatty Acid Free Bsa, supplied by Applygen Technologies, used in various techniques. Bioz Stars score: 94/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Introgen fatty acid free bsa
    The effect of VASP on lipid metabolism in <t>AML12</t> cells. A : AML12 cells were transduced with VASP (VASP-OE) or control (ctl) (empty) vector. B : RT-PCR analysis of lipid metabolism–related genes in AML12 cells. Expression of Gapdh as shown by the ratio of CT value ( n = 3). C : Rate of [1- 14 C]palmitate incorporation into acid-soluble metabolites ( n = 4). D : Oleic acid (either 0.1 or 0.4 mmol/L)-induced accumulation of TG in AML12 cells. <t>BSA</t> (1.3 mmol/L) was used as a control ( n = 3). * P
    Fatty Acid Free Bsa, supplied by Introgen, used in various techniques. Bioz Stars score: 79/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher fatty acid free bsa
    The effect of VASP on lipid metabolism in <t>AML12</t> cells. A : AML12 cells were transduced with VASP (VASP-OE) or control (ctl) (empty) vector. B : RT-PCR analysis of lipid metabolism–related genes in AML12 cells. Expression of Gapdh as shown by the ratio of CT value ( n = 3). C : Rate of [1- 14 C]palmitate incorporation into acid-soluble metabolites ( n = 4). D : Oleic acid (either 0.1 or 0.4 mmol/L)-induced accumulation of TG in AML12 cells. <t>BSA</t> (1.3 mmol/L) was used as a control ( n = 3). * P
    Fatty Acid Free Bsa, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 94/100, based on 207 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GenDEPOT fatty acid free bsa
    The effect of VASP on lipid metabolism in <t>AML12</t> cells. A : AML12 cells were transduced with VASP (VASP-OE) or control (ctl) (empty) vector. B : RT-PCR analysis of lipid metabolism–related genes in AML12 cells. Expression of Gapdh as shown by the ratio of CT value ( n = 3). C : Rate of [1- 14 C]palmitate incorporation into acid-soluble metabolites ( n = 4). D : Oleic acid (either 0.1 or 0.4 mmol/L)-induced accumulation of TG in AML12 cells. <t>BSA</t> (1.3 mmol/L) was used as a control ( n = 3). * P
    Fatty Acid Free Bsa, supplied by GenDEPOT, used in various techniques. Bioz Stars score: 91/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore fatty acid free bovine serum albumin bsa
    Glutathionation in A549 cells supplemented with <t>BSA-conjugated</t> fatty acids (LA, DHA, BSA carrier control). Cells were treated with BSO or <t>GSH</t> in combination with each of the fatty acids for 48 µM BaP for 24 h. Data represents mean normalized intensity ± SEM of at least 30 images per treatment and 30 cells per image. * indicates significant difference from the corresponding control using Tukey test at p
    Fatty Acid Free Bovine Serum Albumin Bsa, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 742 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    BioShop fatty acid free bovine serum albumin bsa
    Glutathionation in A549 cells supplemented with <t>BSA-conjugated</t> fatty acids (LA, DHA, BSA carrier control). Cells were treated with BSO or <t>GSH</t> in combination with each of the fatty acids for 48 µM BaP for 24 h. Data represents mean normalized intensity ± SEM of at least 30 images per treatment and 30 cells per image. * indicates significant difference from the corresponding control using Tukey test at p
    Fatty Acid Free Bovine Serum Albumin Bsa, supplied by BioShop, used in various techniques. Bioz Stars score: 83/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Beijing Solarbio Science fatty acid free bsa
    Glutathionation in A549 cells supplemented with <t>BSA-conjugated</t> fatty acids (LA, DHA, BSA carrier control). Cells were treated with BSO or <t>GSH</t> in combination with each of the fatty acids for 48 µM BaP for 24 h. Data represents mean normalized intensity ± SEM of at least 30 images per treatment and 30 cells per image. * indicates significant difference from the corresponding control using Tukey test at p
    Fatty Acid Free Bsa, supplied by Beijing Solarbio Science, used in various techniques. Bioz Stars score: 94/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Applichem fatty acid free bsa
    Glutathionation in A549 cells supplemented with <t>BSA-conjugated</t> fatty acids (LA, DHA, BSA carrier control). Cells were treated with BSO or <t>GSH</t> in combination with each of the fatty acids for 48 µM BaP for 24 h. Data represents mean normalized intensity ± SEM of at least 30 images per treatment and 30 cells per image. * indicates significant difference from the corresponding control using Tukey test at p
    Fatty Acid Free Bsa, supplied by Applichem, used in various techniques. Bioz Stars score: 83/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Nacalai fatty acid free bsa
    Glutathionation in A549 cells supplemented with <t>BSA-conjugated</t> fatty acids (LA, DHA, BSA carrier control). Cells were treated with BSO or <t>GSH</t> in combination with each of the fatty acids for 48 µM BaP for 24 h. Data represents mean normalized intensity ± SEM of at least 30 images per treatment and 30 cells per image. * indicates significant difference from the corresponding control using Tukey test at p
    Fatty Acid Free Bsa, supplied by Nacalai, used in various techniques. Bioz Stars score: 96/100, based on 26 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Desert Biologicals fatty acid free bsa
    Glutathionation in A549 cells supplemented with <t>BSA-conjugated</t> fatty acids (LA, DHA, BSA carrier control). Cells were treated with BSO or <t>GSH</t> in combination with each of the fatty acids for 48 µM BaP for 24 h. Data represents mean normalized intensity ± SEM of at least 30 images per treatment and 30 cells per image. * indicates significant difference from the corresponding control using Tukey test at p
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    Effect of 24 hour exposure to fatty acids on human BMMSC energy metabolism. A) Palmitate oxidation, B) palmitate uptake, C) glycolysis, and D) glucose oxidation were measured in human <t>BMMSCs</t> that had been treated for 24 hr with either 0.55 mM albumin <t>(BSA</t> group) or 0.55 mM albumin and 0.4 mM palmitate and/or 0.4 mM oleate prior to these metabolism measurements being made. n = 5–8 The graphs indicate which groups were exposed to these different treatments for the 24 hr prior to the metabolism measurements. During each assay all groups were given Krebs buffer supplemented with 5 mM glucose and 0.4 mM palmitate bound to 0.55 mM albumin. In addition, the Krebs buffer was supplemented with either [U- 14 C]glucose, [1– 14 C]palmitate, or [5– 3 H]glucose for the measurement of glucose oxidation, palmitate oxidation and uptake, or glycolysis, respectively. E) The contribution of metabolic pathways to ATP production were calculated from the metabolic rate results. * Significantly different from all groups. Values are shown as mean ± SEM.
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    Boehringer Mannheim fatty acid free bovine serum albumin bsa
    Effect of 24 hour exposure to fatty acids on human BMMSC energy metabolism. A) Palmitate oxidation, B) palmitate uptake, C) glycolysis, and D) glucose oxidation were measured in human <t>BMMSCs</t> that had been treated for 24 hr with either 0.55 mM albumin <t>(BSA</t> group) or 0.55 mM albumin and 0.4 mM palmitate and/or 0.4 mM oleate prior to these metabolism measurements being made. n = 5–8 The graphs indicate which groups were exposed to these different treatments for the 24 hr prior to the metabolism measurements. During each assay all groups were given Krebs buffer supplemented with 5 mM glucose and 0.4 mM palmitate bound to 0.55 mM albumin. In addition, the Krebs buffer was supplemented with either [U- 14 C]glucose, [1– 14 C]palmitate, or [5– 3 H]glucose for the measurement of glucose oxidation, palmitate oxidation and uptake, or glycolysis, respectively. E) The contribution of metabolic pathways to ATP production were calculated from the metabolic rate results. * Significantly different from all groups. Values are shown as mean ± SEM.
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    Effect of 24 hour exposure to fatty acids on human BMMSC energy metabolism. A) Palmitate oxidation, B) palmitate uptake, C) glycolysis, and D) glucose oxidation were measured in human <t>BMMSCs</t> that had been treated for 24 hr with either 0.55 mM albumin <t>(BSA</t> group) or 0.55 mM albumin and 0.4 mM palmitate and/or 0.4 mM oleate prior to these metabolism measurements being made. n = 5–8 The graphs indicate which groups were exposed to these different treatments for the 24 hr prior to the metabolism measurements. During each assay all groups were given Krebs buffer supplemented with 5 mM glucose and 0.4 mM palmitate bound to 0.55 mM albumin. In addition, the Krebs buffer was supplemented with either [U- 14 C]glucose, [1– 14 C]palmitate, or [5– 3 H]glucose for the measurement of glucose oxidation, palmitate oxidation and uptake, or glycolysis, respectively. E) The contribution of metabolic pathways to ATP production were calculated from the metabolic rate results. * Significantly different from all groups. Values are shown as mean ± SEM.
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    Effect of 24 hour exposure to fatty acids on human BMMSC energy metabolism. A) Palmitate oxidation, B) palmitate uptake, C) glycolysis, and D) glucose oxidation were measured in human <t>BMMSCs</t> that had been treated for 24 hr with either 0.55 mM albumin <t>(BSA</t> group) or 0.55 mM albumin and 0.4 mM palmitate and/or 0.4 mM oleate prior to these metabolism measurements being made. n = 5–8 The graphs indicate which groups were exposed to these different treatments for the 24 hr prior to the metabolism measurements. During each assay all groups were given Krebs buffer supplemented with 5 mM glucose and 0.4 mM palmitate bound to 0.55 mM albumin. In addition, the Krebs buffer was supplemented with either [U- 14 C]glucose, [1– 14 C]palmitate, or [5– 3 H]glucose for the measurement of glucose oxidation, palmitate oxidation and uptake, or glycolysis, respectively. E) The contribution of metabolic pathways to ATP production were calculated from the metabolic rate results. * Significantly different from all groups. Values are shown as mean ± SEM.
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    Effect of 24 hour exposure to fatty acids on human BMMSC energy metabolism. A) Palmitate oxidation, B) palmitate uptake, C) glycolysis, and D) glucose oxidation were measured in human <t>BMMSCs</t> that had been treated for 24 hr with either 0.55 mM albumin <t>(BSA</t> group) or 0.55 mM albumin and 0.4 mM palmitate and/or 0.4 mM oleate prior to these metabolism measurements being made. n = 5–8 The graphs indicate which groups were exposed to these different treatments for the 24 hr prior to the metabolism measurements. During each assay all groups were given Krebs buffer supplemented with 5 mM glucose and 0.4 mM palmitate bound to 0.55 mM albumin. In addition, the Krebs buffer was supplemented with either [U- 14 C]glucose, [1– 14 C]palmitate, or [5– 3 H]glucose for the measurement of glucose oxidation, palmitate oxidation and uptake, or glycolysis, respectively. E) The contribution of metabolic pathways to ATP production were calculated from the metabolic rate results. * Significantly different from all groups. Values are shown as mean ± SEM.
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    Image Search Results


    Modulation of S1P homeostasis of K562 cells by sodium butyrate-induced differentiation into erythroblast-like cells. K562 cells were treated with 2 mM sodium butyrate (NaB); 72 hours later, we investigated the modulation of S1P homeostasis. (A) The mRNA levels of GYPA and ALAS were determined using real-time PCR. GAPDH was utilized as an internal control (n = 8/group). (B, C) The expression of CD235a was investigated with a flow cytometer (n = 3/group). (D) C 17 S1P formation assay. NaB-treated or vehicle-treated K562 cells were treated with 10 μM of C 17 sphingosine for 20 minutes. Then, we replaced the supernatant with PBS containing 0.5% BSA and incubated the cells at 37°C for another 20 minutes. Then, the supernatants and cells were collected and used for the C 17 S1P measurements (n = 6/group). (E) The expression of key enzymes in S1P metabolism was determined using real-time PCR. GAPDH was utilized as an internal control (n = 8/group). (F) SK activity assay. The SK activity assay was performed using NaB-treated or vehicle-treated K562 cells (n = 6/group). (G) Reverse transcription PCR was performed using cDNAs prepared from NaB-treated K562 cells (K), HepG2 cells (He), and HUVECs (Hu). (H) The expression of possible S1P transporters was determined using real-time PCR. GAPDH was utilized as an internal control (n = 4/group). (I) Real-time PCR of Band3. GAPDH was utilized as an internal control (n = 8/group). (J) Western blot of Band3 with membranous protein. The whole cell lysate of RBCs (2 μg) was placed as a positive control. Pan-cadherin was utilized as an internal control (n = 3/group).

    Journal: PLoS ONE

    Article Title: Involvement of Band3 in the efflux of sphingosine 1-phosphate from erythrocytes

    doi: 10.1371/journal.pone.0177543

    Figure Lengend Snippet: Modulation of S1P homeostasis of K562 cells by sodium butyrate-induced differentiation into erythroblast-like cells. K562 cells were treated with 2 mM sodium butyrate (NaB); 72 hours later, we investigated the modulation of S1P homeostasis. (A) The mRNA levels of GYPA and ALAS were determined using real-time PCR. GAPDH was utilized as an internal control (n = 8/group). (B, C) The expression of CD235a was investigated with a flow cytometer (n = 3/group). (D) C 17 S1P formation assay. NaB-treated or vehicle-treated K562 cells were treated with 10 μM of C 17 sphingosine for 20 minutes. Then, we replaced the supernatant with PBS containing 0.5% BSA and incubated the cells at 37°C for another 20 minutes. Then, the supernatants and cells were collected and used for the C 17 S1P measurements (n = 6/group). (E) The expression of key enzymes in S1P metabolism was determined using real-time PCR. GAPDH was utilized as an internal control (n = 8/group). (F) SK activity assay. The SK activity assay was performed using NaB-treated or vehicle-treated K562 cells (n = 6/group). (G) Reverse transcription PCR was performed using cDNAs prepared from NaB-treated K562 cells (K), HepG2 cells (He), and HUVECs (Hu). (H) The expression of possible S1P transporters was determined using real-time PCR. GAPDH was utilized as an internal control (n = 4/group). (I) Real-time PCR of Band3. GAPDH was utilized as an internal control (n = 8/group). (J) Western blot of Band3 with membranous protein. The whole cell lysate of RBCs (2 μg) was placed as a positive control. Pan-cadherin was utilized as an internal control (n = 3/group).

    Article Snippet: Then, we replaced the supernatant with PBS containing 0.5% fatty acid-free BSA (A8806; Sigma-Aldrich Co.) and incubated the cells at 37°C for another 20 minutes.

    Techniques: Real-time Polymerase Chain Reaction, Expressing, Flow Cytometry, Cytometry, Tube Formation Assay, Incubation, Activity Assay, Polymerase Chain Reaction, Western Blot, Positive Control

    Effects of FBS, BSA, and essential fatty acid-free BSA on retention of the glycocalyx. Immunofluorescence images ( A ) and coverage analysis ( B ) show the effects of FBS, BSA, and essential fatty acid-free BSA on CS. The average S1P content was 100, 87,

    Journal: American Journal of Physiology - Heart and Circulatory Physiology

    Article Title: Sphingosine-1-phosphate protects endothelial glycocalyx by inhibiting syndecan-1 shedding

    doi: 10.1152/ajpheart.00687.2013

    Figure Lengend Snippet: Effects of FBS, BSA, and essential fatty acid-free BSA on retention of the glycocalyx. Immunofluorescence images ( A ) and coverage analysis ( B ) show the effects of FBS, BSA, and essential fatty acid-free BSA on CS. The average S1P content was 100, 87,

    Article Snippet: The effects of plasma protein on CS were further determined by replacing the 5% FBS and 0.5% BSA with 5% FBS alone, 0.5% BSA alone, or 0.5% essential fatty acid-free BSA (A6003, Sigma).

    Techniques: Immunofluorescence

    G2A-dependent chemotaxis to LPC. Equal numbers (2 × 10 5 ) of WT and G2A shRNA or control (CTR shRNA ) DO11.10 cells were washed three times with SFM containing 0.1% FAF-BSA, mixed, and added to the upper chamber of a 24-well plate with 5.0-μm pore size polycarbonate filters (Costar). LPC ( A ) or SDF1-α ( B ) was added to the lower chamber. After a 2-h incubation at 37°C in an 8% CO 2 incubator, transmigrated WT and EGFP-positive siRNA-transduced cells were counted by fluorescence-activated cell sorting. The results are representative of four independent experiments.

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

    Article Title: T cell chemotaxis to lysophosphatidylcholine through the G2A receptor

    doi: 10.1073/pnas.2536801100

    Figure Lengend Snippet: G2A-dependent chemotaxis to LPC. Equal numbers (2 × 10 5 ) of WT and G2A shRNA or control (CTR shRNA ) DO11.10 cells were washed three times with SFM containing 0.1% FAF-BSA, mixed, and added to the upper chamber of a 24-well plate with 5.0-μm pore size polycarbonate filters (Costar). LPC ( A ) or SDF1-α ( B ) was added to the lower chamber. After a 2-h incubation at 37°C in an 8% CO 2 incubator, transmigrated WT and EGFP-positive siRNA-transduced cells were counted by fluorescence-activated cell sorting. The results are representative of four independent experiments.

    Article Snippet: Cells (5 × 104 ) were resuspended in serum-free medium (SFM) containing RPMI medium 1640 with 0.1% fatty acid-free BSA (FAF-BSA) (Sigma, no. A8806-5G) and 25 mM Hepes buffer, pH 7.4.

    Techniques: Chemotaxis Assay, shRNA, Incubation, Fluorescence, FACS

    Time course of SAA and other inflammatory mediators in mesangial cells stimulated with AGE. Cells were exposed to (i) control (no addition) and (ii) AGE (AGE-BSA, 300 µg/mL). Expression of mRNA was determined by real-time RT–PCR and displayed

    Journal: Nephrology Dialysis Transplantation

    Article Title: Glomerular cell death and inflammation with high-protein diet and diabetes

    doi: 10.1093/ndt/gfs579

    Figure Lengend Snippet: Time course of SAA and other inflammatory mediators in mesangial cells stimulated with AGE. Cells were exposed to (i) control (no addition) and (ii) AGE (AGE-BSA, 300 µg/mL). Expression of mRNA was determined by real-time RT–PCR and displayed

    Article Snippet: To make AGE-BSA, fatty acid-free fraction IV BSA (Sigma) was incubated with 0.5 M glucose for 45 days at 37°C.

    Techniques: Expressing, Quantitative RT-PCR

    SAA and other inflammatory mediators expressed in podocytes exposed to AGE and inhibition by antibody to RAGE. Cells were exposed to (i) control and (ii) AGE-bovine serum albumin (AGE-BSA, 300 µg/mL), with and without the anti-RAGE antibody or

    Journal: Nephrology Dialysis Transplantation

    Article Title: Glomerular cell death and inflammation with high-protein diet and diabetes

    doi: 10.1093/ndt/gfs579

    Figure Lengend Snippet: SAA and other inflammatory mediators expressed in podocytes exposed to AGE and inhibition by antibody to RAGE. Cells were exposed to (i) control and (ii) AGE-bovine serum albumin (AGE-BSA, 300 µg/mL), with and without the anti-RAGE antibody or

    Article Snippet: To make AGE-BSA, fatty acid-free fraction IV BSA (Sigma) was incubated with 0.5 M glucose for 45 days at 37°C.

    Techniques: Inhibition

    Palmitate induced oxidative stress in INS-1 cells. INS-1 cells were treated with control medium, FFA-free BSA (0.5 mol/ml), or indicated concentration of palmitate 24 h or 0.5 mM of palmitate for indicated times. The results showed that palmitate increased the level of ROS in INS-1 cells, which was measured by flow cytometry (B, D) and fluorescence microscopy (A, C). Data are expressed as means ± SEM of 3 independent experiments; * P

    Journal: PLoS ONE

    Article Title: Interleukin-22 Alleviated Palmitate-Induced Endoplasmic Reticulum Stress in INS-1 Cells through Activation of Autophagy

    doi: 10.1371/journal.pone.0146818

    Figure Lengend Snippet: Palmitate induced oxidative stress in INS-1 cells. INS-1 cells were treated with control medium, FFA-free BSA (0.5 mol/ml), or indicated concentration of palmitate 24 h or 0.5 mM of palmitate for indicated times. The results showed that palmitate increased the level of ROS in INS-1 cells, which was measured by flow cytometry (B, D) and fluorescence microscopy (A, C). Data are expressed as means ± SEM of 3 independent experiments; * P

    Article Snippet: A 5% (w/v) solution of FFA-free bovine serum albumin (BSA) (Sigma-Aldrich, Milano, Italy) was prepared in serum-free RPMI medium.

    Techniques: Concentration Assay, Flow Cytometry, Cytometry, Fluorescence, Microscopy

    Determination of neutral lipid accumulation by ORO staining in OA-loaded HepaRG cells. HepaRG cells were incubated for 48 hours in HepaRG treatment medium alone (CON) or containing 0.7% FF-BSA, either alone or complexed with 0.33 mM, 0.66 mM, or 1 mM OA. Treatment medium was replaced once after 24 hours. (A) ORO was used to stain neutral lipids, and cells were photographed under phase-contrast optics at 200X magnification. Neutral lipids are stained red, hepatocyte nuclei are purple/blue. (B) ORO was extracted from the cells and absorbance measured at 510 nm. Each bar represents the mean ± SEM (n=4 wells per group, derived from combining the data from 2 independent HepaRG experiments with duplicate treatments). *Significantly different from vehicle control (FF-BSA), P

    Journal: Toxicology and applied pharmacology

    Article Title: Farnesol induces fatty acid oxidation and decreases triglyceride accumulation in steatotic HepaRG cells

    doi: 10.1016/j.taap.2019.01.003

    Figure Lengend Snippet: Determination of neutral lipid accumulation by ORO staining in OA-loaded HepaRG cells. HepaRG cells were incubated for 48 hours in HepaRG treatment medium alone (CON) or containing 0.7% FF-BSA, either alone or complexed with 0.33 mM, 0.66 mM, or 1 mM OA. Treatment medium was replaced once after 24 hours. (A) ORO was used to stain neutral lipids, and cells were photographed under phase-contrast optics at 200X magnification. Neutral lipids are stained red, hepatocyte nuclei are purple/blue. (B) ORO was extracted from the cells and absorbance measured at 510 nm. Each bar represents the mean ± SEM (n=4 wells per group, derived from combining the data from 2 independent HepaRG experiments with duplicate treatments). *Significantly different from vehicle control (FF-BSA), P

    Article Snippet: Trans, trans -FOH, dimethyl sulfoxide (DMSO), 6-(4-chlorophenyl)imidazo[2,1-b][1,3]thiazole-5-carbaldehyde O-(3,4-dichlorobenzyl)oxime (CITCO), 3-[2-[2-chloro-4-[[3-(2,6-dichlorophenyl)-5-(1-methylethyl)-4 isoxazolyl]methoxy]phenyl]ethenyl]benzoic acid (GW4064), OA, hematoxylin, Oil Red O (ORO), 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), triamcinolone acetonide, Triton X-100, and fatty acid-free bovine serum albumin (FF-BSA) were purchased from Sigma-Aldrich (St. Louis, MO).

    Techniques: Staining, Incubation, Derivative Assay

    Effects of FOH on HepaRG viability and TG content. (A) HepaRG cells were cultured for 24 or 48 hours in HepaRG treatment medium containing either vehicle control [FFBSA+0.1% EtOH (EtOH)] alone or FF-BSA complexed with 10 – 1000 μM FOH. Cell viability was determined using the MTT assay, as described in Materials and Methods. Each bar represents the mean MTT formazan absorbance ± SEM from three independent experiments (n=3). Values were normalized to those of the EtOH-treated group. (B) HepaRG cells were pre-treated for 24 hours in HepaRG treatment medium containing FF-BSA, either alone or complexed with 0.66 mM OA. The cells were then incubated for 48 hours in pre-treatment medium (i.e., FF-BSA or OA) containing either 0.1% EtOH or 100 μM FOH and harvested for TG measurement, as described in Materials and Methods. The amount of TG was normalized to protein content for each sample. Each bar is the mean ± SEM from 3 independent experiments (n=3). ***Significantly different from FF-BSA:EtOH group, P

    Journal: Toxicology and applied pharmacology

    Article Title: Farnesol induces fatty acid oxidation and decreases triglyceride accumulation in steatotic HepaRG cells

    doi: 10.1016/j.taap.2019.01.003

    Figure Lengend Snippet: Effects of FOH on HepaRG viability and TG content. (A) HepaRG cells were cultured for 24 or 48 hours in HepaRG treatment medium containing either vehicle control [FFBSA+0.1% EtOH (EtOH)] alone or FF-BSA complexed with 10 – 1000 μM FOH. Cell viability was determined using the MTT assay, as described in Materials and Methods. Each bar represents the mean MTT formazan absorbance ± SEM from three independent experiments (n=3). Values were normalized to those of the EtOH-treated group. (B) HepaRG cells were pre-treated for 24 hours in HepaRG treatment medium containing FF-BSA, either alone or complexed with 0.66 mM OA. The cells were then incubated for 48 hours in pre-treatment medium (i.e., FF-BSA or OA) containing either 0.1% EtOH or 100 μM FOH and harvested for TG measurement, as described in Materials and Methods. The amount of TG was normalized to protein content for each sample. Each bar is the mean ± SEM from 3 independent experiments (n=3). ***Significantly different from FF-BSA:EtOH group, P

    Article Snippet: Trans, trans -FOH, dimethyl sulfoxide (DMSO), 6-(4-chlorophenyl)imidazo[2,1-b][1,3]thiazole-5-carbaldehyde O-(3,4-dichlorobenzyl)oxime (CITCO), 3-[2-[2-chloro-4-[[3-(2,6-dichlorophenyl)-5-(1-methylethyl)-4 isoxazolyl]methoxy]phenyl]ethenyl]benzoic acid (GW4064), OA, hematoxylin, Oil Red O (ORO), 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), triamcinolone acetonide, Triton X-100, and fatty acid-free bovine serum albumin (FF-BSA) were purchased from Sigma-Aldrich (St. Louis, MO).

    Techniques: Cell Culture, MTT Assay, Incubation

    Calibration curve of carbonylated proteins for quantitation of carbonyl indexes by dot-blot. ( A ). 2 μL of DNPH derivatized bovine serum albumin (BSA; 100 ng/μL) from standard working solutions were spotted by triplicate on PVDF membranes. ( B ). Mean from three calibration curves analyzed in two different days. The calibration curve was derived from reproducibility assay.

    Journal: International Journal of Molecular Sciences

    Article Title: Sickle Cell Trait Induces Oxidative Damage on Plasmodium falciparum Proteome at Erythrocyte Stages

    doi: 10.3390/ijms20225769

    Figure Lengend Snippet: Calibration curve of carbonylated proteins for quantitation of carbonyl indexes by dot-blot. ( A ). 2 μL of DNPH derivatized bovine serum albumin (BSA; 100 ng/μL) from standard working solutions were spotted by triplicate on PVDF membranes. ( B ). Mean from three calibration curves analyzed in two different days. The calibration curve was derived from reproducibility assay.

    Article Snippet: For this purpose, an aliquot of the BSA fatty acid-free solution (5 μg/μL, Sigma) was derivatized with a DNPH probe, and their carbonyl index was measured in alkaline medium at 450 nm [ ].

    Techniques: Quantitation Assay, Dot Blot, Derivative Assay

    4BA inhibits dark adaptation in 11-cis-retinal–regenerated WT-HEK293S cells. Giant cells were regenerated with 50 μM 11-cis-retinal and cell surface area–normalized R 2 charge motion obtained for the first bleach cycle (Control, c). 4BA complexed to FAF-BSA was then perfused through the chamber and the amount of R 2 charge motion obtained in successive bleach cycles determined (1Æ5). Recovery time between bleach cycles was 10 min in all cases and flash stimulation was 500 nm. After the fifth bleach cycle, 4BA was washed out of the chamber and the recovery of R 2 charge determined (washout, w). Control and washout conditions were not statistically different. The decay of R 2 charge motion in 4BA (0.5 mM) was reliably fit with a single exponential model with a decay constant of 0.45 ± 0.01 cycles ( P

    Journal: The Journal of General Physiology

    Article Title: HEK293S Cells Have Functional Retinoid Processing Machinery

    doi: 10.1085/jgp.20018495

    Figure Lengend Snippet: 4BA inhibits dark adaptation in 11-cis-retinal–regenerated WT-HEK293S cells. Giant cells were regenerated with 50 μM 11-cis-retinal and cell surface area–normalized R 2 charge motion obtained for the first bleach cycle (Control, c). 4BA complexed to FAF-BSA was then perfused through the chamber and the amount of R 2 charge motion obtained in successive bleach cycles determined (1Æ5). Recovery time between bleach cycles was 10 min in all cases and flash stimulation was 500 nm. After the fifth bleach cycle, 4BA was washed out of the chamber and the recovery of R 2 charge determined (washout, w). Control and washout conditions were not statistically different. The decay of R 2 charge motion in 4BA (0.5 mM) was reliably fit with a single exponential model with a decay constant of 0.45 ± 0.01 cycles ( P

    Article Snippet: All-trans-retinal, 9-cis-retinal, 13-cis-retinal, all-trans-retinol (Vitamin A), α-D-tocopherol (Vitamin E), and essentially fatty acid–free (FAF) BSA were obtained from Sigma-Aldrich.

    Techniques:

    ERC evidence for retinoid conversions in giant WT-HEK293S cells. Cells were loaded with 50 μM chromophore complexed to FAF-BSA in all cases. ERCs were obtained upon the first 500-nm flash from cells regenerated for 40 min at room temperature in darkness (A and C) or overnight at 4°C in darkness (B and D). Each panel is from a single cell. Cells were regenerated with all-trans-retinal (A and B) or Vitamin A (C and D). Cells are representative of larger populations of cells examined (all-trans-retinal, 40 min: n = 43, overnight at 4°C: n = 7; Vitamin A, 40 min: n = 2 cells, overnight at 4°C: n = 23 cells).

    Journal: The Journal of General Physiology

    Article Title: HEK293S Cells Have Functional Retinoid Processing Machinery

    doi: 10.1085/jgp.20018495

    Figure Lengend Snippet: ERC evidence for retinoid conversions in giant WT-HEK293S cells. Cells were loaded with 50 μM chromophore complexed to FAF-BSA in all cases. ERCs were obtained upon the first 500-nm flash from cells regenerated for 40 min at room temperature in darkness (A and C) or overnight at 4°C in darkness (B and D). Each panel is from a single cell. Cells were regenerated with all-trans-retinal (A and B) or Vitamin A (C and D). Cells are representative of larger populations of cells examined (all-trans-retinal, 40 min: n = 43, overnight at 4°C: n = 7; Vitamin A, 40 min: n = 2 cells, overnight at 4°C: n = 23 cells).

    Article Snippet: All-trans-retinal, 9-cis-retinal, 13-cis-retinal, all-trans-retinol (Vitamin A), α-D-tocopherol (Vitamin E), and essentially fatty acid–free (FAF) BSA were obtained from Sigma-Aldrich.

    Techniques:

    Lack of bulk BSA uptake into single or giant WT-HEK293S cells. Single (A and B) or giant PEG-fused (C and D) WT-HEK293S cells were exposed to regeneration buffer containing 1.9% (wt/vol) FAF-BSA plus 0.1% FITC-BSA. Hoffman contrast images of representative fields of single (A) or PEG-fused (C) WT-HEK293S cells are shown beside fluorescence images from the same respective fields (B and D). Arrows in A and B indicate healthy single cells and arrowheads indicate unhealthy single cells and single cells or debris taking up FITC-BSA. Scale marker is 20 μm in all fields and approximates the size of a single unfused HEK293S cell.

    Journal: The Journal of General Physiology

    Article Title: HEK293S Cells Have Functional Retinoid Processing Machinery

    doi: 10.1085/jgp.20018495

    Figure Lengend Snippet: Lack of bulk BSA uptake into single or giant WT-HEK293S cells. Single (A and B) or giant PEG-fused (C and D) WT-HEK293S cells were exposed to regeneration buffer containing 1.9% (wt/vol) FAF-BSA plus 0.1% FITC-BSA. Hoffman contrast images of representative fields of single (A) or PEG-fused (C) WT-HEK293S cells are shown beside fluorescence images from the same respective fields (B and D). Arrows in A and B indicate healthy single cells and arrowheads indicate unhealthy single cells and single cells or debris taking up FITC-BSA. Scale marker is 20 μm in all fields and approximates the size of a single unfused HEK293S cell.

    Article Snippet: All-trans-retinal, 9-cis-retinal, 13-cis-retinal, all-trans-retinol (Vitamin A), α-D-tocopherol (Vitamin E), and essentially fatty acid–free (FAF) BSA were obtained from Sigma-Aldrich.

    Techniques: Fluorescence, Marker

    ERC signals from WT-HEK293S cells regenerated with different cis-retinaldehydes. Fused WT-HEK293S giant cells were loaded with 25 μM 11-cis-retinal or 9-cis-retinal or 50 μM 13-cis-retinal complexed to FAF-BSA. ERC signals on the first 500-nm flash during the primary bleaching extinction (A, C, and E) and secondary bleaching extinctions (B, D, and F) are shown for 11-cis-retinal– (A and B), 9-cis-retinal– (C and D), and 13-cis-retinal– (E and F) loaded representative cells. Membrane capacitances of the cells are indicated. The arrow indicates the timing of the flash stimulus. Responses from each single cell are representative of larger populations of cells regenerated with 11-cis-retinal ( n = 54), 9-cis-retinal ( n = 5), and 13-cis-retinal ( n = 4).

    Journal: The Journal of General Physiology

    Article Title: HEK293S Cells Have Functional Retinoid Processing Machinery

    doi: 10.1085/jgp.20018495

    Figure Lengend Snippet: ERC signals from WT-HEK293S cells regenerated with different cis-retinaldehydes. Fused WT-HEK293S giant cells were loaded with 25 μM 11-cis-retinal or 9-cis-retinal or 50 μM 13-cis-retinal complexed to FAF-BSA. ERC signals on the first 500-nm flash during the primary bleaching extinction (A, C, and E) and secondary bleaching extinctions (B, D, and F) are shown for 11-cis-retinal– (A and B), 9-cis-retinal– (C and D), and 13-cis-retinal– (E and F) loaded representative cells. Membrane capacitances of the cells are indicated. The arrow indicates the timing of the flash stimulus. Responses from each single cell are representative of larger populations of cells regenerated with 11-cis-retinal ( n = 54), 9-cis-retinal ( n = 5), and 13-cis-retinal ( n = 4).

    Article Snippet: All-trans-retinal, 9-cis-retinal, 13-cis-retinal, all-trans-retinol (Vitamin A), α-D-tocopherol (Vitamin E), and essentially fatty acid–free (FAF) BSA were obtained from Sigma-Aldrich.

    Techniques:

    p21-dependent inhibition of LPA-induced cell proliferation by TGFβ. A . TGFβ inhibits LPA-afforded cell proliferation. MDA-MB-231 and Caov-3 cells in 6-well plates were incubated with LPA (10 µM) or vehicle (BSA) in the presence

    Journal: Molecular Cancer Research

    Article Title: Lysophosphatidic acid-induced p21Waf1 expression mediates the cytostatic response of breast and ovarian cancer cells to transforming growth factor beta

    doi: 10.1158/1541-7786.MCR-11-0340

    Figure Lengend Snippet: p21-dependent inhibition of LPA-induced cell proliferation by TGFβ. A . TGFβ inhibits LPA-afforded cell proliferation. MDA-MB-231 and Caov-3 cells in 6-well plates were incubated with LPA (10 µM) or vehicle (BSA) in the presence

    Article Snippet: Prior to use, LPA and S1P were dissolved in PBS containing 0.5% fatty acid-free bovine serum albumin (BSA) obtained from Roche (Indianapolis, IN).

    Techniques: Inhibition, Multiple Displacement Amplification, Incubation

    Effect of diabetes on cholesterol efflux to apoA1 or HDL and regulation of cholesterol transporters by Ager in primary murine BMDMs. Primary BMDMs were retrieved from nondiabetic and diabetic WT (2 months of hyperglycemia) and Ager −/− mice and were cultured in concentrations of glucose consistent with the glycemic state of the mice from which they were retrieved. BMDMs were labeled with 3 H-cholesterol and treated with acetylated LDL and fatty acid–free BSA (1%) for 24 h. A and B : To mediate cholesterol efflux, cells were treated with apoA1 (5 μg/mL) ( A ) or HDL (100 μg/mL) ( B ) for 6 h, and supernatant was collected. Percent cholesterol efflux is the radioactivity in the supernatant divided by the sum of the radioactivity measured in the supernatant and the cell lysates from n ≥ 5 mice/group. C and D : BMDMs were retrieved from WT or Ager −/− mice and subjected to quantitative real-time PCR for detection of Abca1 mRNA transcript ( C ) and ABCA1 protein ( D ) analysis ( n = 3 mice/group). E and F : BMDMs were prepared as in C and D and assessed for levels of Abcg1 mRNA transcripts ( E ) and ABCG1 protein ( F ) ( n = 3 mice/group). Error bars represent mean ± SEM. hr, hour; NS, not statistically significant.

    Journal: Diabetes

    Article Title: RAGE Suppresses ABCG1-Mediated Macrophage Cholesterol Efflux in Diabetes

    doi: 10.2337/db15-0575

    Figure Lengend Snippet: Effect of diabetes on cholesterol efflux to apoA1 or HDL and regulation of cholesterol transporters by Ager in primary murine BMDMs. Primary BMDMs were retrieved from nondiabetic and diabetic WT (2 months of hyperglycemia) and Ager −/− mice and were cultured in concentrations of glucose consistent with the glycemic state of the mice from which they were retrieved. BMDMs were labeled with 3 H-cholesterol and treated with acetylated LDL and fatty acid–free BSA (1%) for 24 h. A and B : To mediate cholesterol efflux, cells were treated with apoA1 (5 μg/mL) ( A ) or HDL (100 μg/mL) ( B ) for 6 h, and supernatant was collected. Percent cholesterol efflux is the radioactivity in the supernatant divided by the sum of the radioactivity measured in the supernatant and the cell lysates from n ≥ 5 mice/group. C and D : BMDMs were retrieved from WT or Ager −/− mice and subjected to quantitative real-time PCR for detection of Abca1 mRNA transcript ( C ) and ABCA1 protein ( D ) analysis ( n = 3 mice/group). E and F : BMDMs were prepared as in C and D and assessed for levels of Abcg1 mRNA transcripts ( E ) and ABCG1 protein ( F ) ( n = 3 mice/group). Error bars represent mean ± SEM. hr, hour; NS, not statistically significant.

    Article Snippet: Reagents The following materials were purchased: apolipoprotein A1 (apoA1), HDL, and acetylated LDL (Biomedical Technologies, Inc.); fatty acid–free BSA (Equitech-Bio); T 0901317 (Tocris Bioscience); human plasma LDL (Sigma-Aldrich); assays for measurements of HDL, total cholesterol, and triglycerides (Wako Diagnostics); rosiglitazone (Sigma-Aldrich); and U0126 and PD98509 (Cell Signaling).

    Techniques: Mouse Assay, Cell Culture, Labeling, Radioactivity, Real-time Polymerase Chain Reaction

    C-terminus of ATP11C is required for inhibition of flippase activities by PKC activation. a – c Ba/F3 cells stably expressing HA-tagged ATP11A, ATP11C, and ATP11AAC chimeric proteins were treated with vehicle alone ( a ), 400 nM PMA ( b ), or 1 μM A23187 in the presence of CaCl 2 ( c ) for 15 min. The cells were fixed and immunostained with anti-HA and anti-ATP1A1 antibodies, followed by incubation with Cy3-conjugated anti-rat and Alexa Fluor 488-conjugated anti-rabbit secondary antibodies. Scale bars, 5 μm. d – f Ba/F3 cells were treated with vehicle alone (white bars), 400 nM PMA (black bars), or 400 nM PMA and 2 μM of BIM-1 (gray bars) for 15 min. The cells were then washed with flippase assay buffer and incubated at 15 °C for 5 min with NBD-PS ( d ), or 15 min with NBD-PE ( e ) or NBD-PC ( f ). After extraction with fatty acid-free BSA, the residual fluorescence intensity associated with the cells was determined by flow cytometry. Fold increase of NBD-lipids uptake is shown relative to that in Mock-treated parental Ba/F3 cells (−). Graphs display averages from three independent experiments ± SD. ** p

    Journal: Nature Communications

    Article Title: Phospholipid flippase ATP11C is endocytosed and downregulated following Ca2+-mediated protein kinase C activation

    doi: 10.1038/s41467-017-01338-1

    Figure Lengend Snippet: C-terminus of ATP11C is required for inhibition of flippase activities by PKC activation. a – c Ba/F3 cells stably expressing HA-tagged ATP11A, ATP11C, and ATP11AAC chimeric proteins were treated with vehicle alone ( a ), 400 nM PMA ( b ), or 1 μM A23187 in the presence of CaCl 2 ( c ) for 15 min. The cells were fixed and immunostained with anti-HA and anti-ATP1A1 antibodies, followed by incubation with Cy3-conjugated anti-rat and Alexa Fluor 488-conjugated anti-rabbit secondary antibodies. Scale bars, 5 μm. d – f Ba/F3 cells were treated with vehicle alone (white bars), 400 nM PMA (black bars), or 400 nM PMA and 2 μM of BIM-1 (gray bars) for 15 min. The cells were then washed with flippase assay buffer and incubated at 15 °C for 5 min with NBD-PS ( d ), or 15 min with NBD-PE ( e ) or NBD-PC ( f ). After extraction with fatty acid-free BSA, the residual fluorescence intensity associated with the cells was determined by flow cytometry. Fold increase of NBD-lipids uptake is shown relative to that in Mock-treated parental Ba/F3 cells (−). Graphs display averages from three independent experiments ± SD. ** p

    Article Snippet: At each time point, 200 μl of the cell suspension was mixed with 200 μl of ice-cold HBSS-glucose containing 5% fatty acid-free BSA (Wako) to extract NBD-lipids incorporated into the exoplasmic leaflet of the plasma membrane, as well as unincorporated lipids.

    Techniques: Inhibition, Activation Assay, Stable Transfection, Expressing, Incubation, Fluorescence, Flow Cytometry, Cytometry

    ATP11C is endocytosed by treatment with phorbol 12-myristate 13-acetate (PMA) and increasing cytosolic Ca 2+ . a HeLa cells stably expressing C-terminally HA-tagged ATP11A, and ATP11C were treated for 15 min at 37 °C with vehicle alone (Mock); with either 400 nM of PMA (PMA) or PMA and 2 μM of BIM-1 (PMA + BIM); or with 1 μM A23187 in the presence of either 1.8 mM of CaCl 2 (A23187) or 1.8 mM CaCl 2 and 2 μM BIM-1 (A23187 + BIM). The cells were fixed and immunostained with anti-HA antibody, followed by Cy3-conjugated anti-rat secondary antibody. See Supplementary Fig. 1 and Supplementary Movie 1 . b Cell-surface expression levels of ATP11A and ATP11C following treatment with PMA or A23187 and CaCl 2 were analyzed after surface biotinylation. Proteins precipitated with streptavidin-agarose beads were subjected to immunoblot analysis (left panels, biotinylated). 15% of the input of the biotinylation reaction was loaded in each lane (right panels, total lysate). Expression of ATP11A and ATP11C proteins was analyzed by immunoblotting with anti-HA and anti-ATP1A1 (as an internal control) antibodies. c HeLa cells stably expressing HA-tagged ATP11A and ATP11C, and parental cells (−) were treated with vehicle alone (white bars), 400 nM of PMA (black bars), or 400 nM of PMA and 2 μM of BIM-1, simultaneously (gray bars) for 15 min. The cells were then washed with flippase assay buffer and incubated with NBD-PS at 15 °C for 5 min. After extraction with fatty acid-free BSA, the residual fluorescence intensity associated with the cells was determined by flow cytometry. Fold increase of NBD-PS uptake is shown relative to that in Mock-treated parental HeLa cells (−). Graph displays averages from four independent experiments ± SD. *** p

    Journal: Nature Communications

    Article Title: Phospholipid flippase ATP11C is endocytosed and downregulated following Ca2+-mediated protein kinase C activation

    doi: 10.1038/s41467-017-01338-1

    Figure Lengend Snippet: ATP11C is endocytosed by treatment with phorbol 12-myristate 13-acetate (PMA) and increasing cytosolic Ca 2+ . a HeLa cells stably expressing C-terminally HA-tagged ATP11A, and ATP11C were treated for 15 min at 37 °C with vehicle alone (Mock); with either 400 nM of PMA (PMA) or PMA and 2 μM of BIM-1 (PMA + BIM); or with 1 μM A23187 in the presence of either 1.8 mM of CaCl 2 (A23187) or 1.8 mM CaCl 2 and 2 μM BIM-1 (A23187 + BIM). The cells were fixed and immunostained with anti-HA antibody, followed by Cy3-conjugated anti-rat secondary antibody. See Supplementary Fig. 1 and Supplementary Movie 1 . b Cell-surface expression levels of ATP11A and ATP11C following treatment with PMA or A23187 and CaCl 2 were analyzed after surface biotinylation. Proteins precipitated with streptavidin-agarose beads were subjected to immunoblot analysis (left panels, biotinylated). 15% of the input of the biotinylation reaction was loaded in each lane (right panels, total lysate). Expression of ATP11A and ATP11C proteins was analyzed by immunoblotting with anti-HA and anti-ATP1A1 (as an internal control) antibodies. c HeLa cells stably expressing HA-tagged ATP11A and ATP11C, and parental cells (−) were treated with vehicle alone (white bars), 400 nM of PMA (black bars), or 400 nM of PMA and 2 μM of BIM-1, simultaneously (gray bars) for 15 min. The cells were then washed with flippase assay buffer and incubated with NBD-PS at 15 °C for 5 min. After extraction with fatty acid-free BSA, the residual fluorescence intensity associated with the cells was determined by flow cytometry. Fold increase of NBD-PS uptake is shown relative to that in Mock-treated parental HeLa cells (−). Graph displays averages from four independent experiments ± SD. *** p

    Article Snippet: At each time point, 200 μl of the cell suspension was mixed with 200 μl of ice-cold HBSS-glucose containing 5% fatty acid-free BSA (Wako) to extract NBD-lipids incorporated into the exoplasmic leaflet of the plasma membrane, as well as unincorporated lipids.

    Techniques: Stable Transfection, Expressing, Incubation, Fluorescence, Flow Cytometry, Cytometry

    Analyses of flippase activity in different human cancer cell types A. Flippase activity assay: Indicated cell types were incubated with NBD-PS for indicated time periods and subjected to BSA extraction and sodium dithionite treatment. % nonextractable NBD-PS (after BSA extraction and sodium dithionite treatment) represents internalized NBD-PS, indicative of flippase activity (Schwann ◊, U87ΔEGFR-Luc □, H1299 Δ, MDA-MB-231 ○, MDA-MB-231-Luc-D3H2LN ◆, Gli36 ▲, U373 ●) B. NBD PS incorporation rate. C. NBD PS incorporation at 15 minutes time point.

    Journal: Oncotarget

    Article Title: Variation in human cancer cell external phosphatidylserine is regulated by flippase activity and intracellular calcium

    doi:

    Figure Lengend Snippet: Analyses of flippase activity in different human cancer cell types A. Flippase activity assay: Indicated cell types were incubated with NBD-PS for indicated time periods and subjected to BSA extraction and sodium dithionite treatment. % nonextractable NBD-PS (after BSA extraction and sodium dithionite treatment) represents internalized NBD-PS, indicative of flippase activity (Schwann ◊, U87ΔEGFR-Luc □, H1299 Δ, MDA-MB-231 ○, MDA-MB-231-Luc-D3H2LN ◆, Gli36 ▲, U373 ●) B. NBD PS incorporation rate. C. NBD PS incorporation at 15 minutes time point.

    Article Snippet: The remaining half was spun down to remove non inserted NBD-PS and subjected to BSA extraction of NBD-PS from the outer leaflet by adding 3% fatty acid free BSA (MP biomedicals) in flippase assay buffer.

    Techniques: Activity Assay, Incubation, Multiple Displacement Amplification

    Inhibition of flippase activity by NEM reveals involvement of flippase activity in the regulation of surface PS A . Low surface PS cells were either treated with NEM or left untreated, incubated with NBD-PS for indicated time periods and subjected to BSA extraction and sodium dithionite treatment. % nonextractable NBD-PS (after BSA extraction and sodium dithionite treatment) represents internalized NBD-PS, indicative of flippase activity B . Flippase activity was inhibited by use of NEM in low surface PS cell lines and surface PS levels were measured by annexin V FITC binding assay, by flow cytometry. The graph shows annexin V FITC fold change compared to mock treated cells. C . High surface PS cells were either treated with NEM or left untreated and incubated with NBD-PS for indicated time periods and subjected to BSA extraction and sodium dithionite treatment. % nonextractable NBD-PS (after BSA extraction and sodium dithionite treatment) represents internalized NBD-PS, indicative of flippase activity D . Flippase activity was inhibited by use of NEM in the high surface PS cell lines and surface PS levels were measured by annexin V FITC binding assay by flow cytometry. The graph shows annexin V FITC fold change compared to mock treated cells.

    Journal: Oncotarget

    Article Title: Variation in human cancer cell external phosphatidylserine is regulated by flippase activity and intracellular calcium

    doi:

    Figure Lengend Snippet: Inhibition of flippase activity by NEM reveals involvement of flippase activity in the regulation of surface PS A . Low surface PS cells were either treated with NEM or left untreated, incubated with NBD-PS for indicated time periods and subjected to BSA extraction and sodium dithionite treatment. % nonextractable NBD-PS (after BSA extraction and sodium dithionite treatment) represents internalized NBD-PS, indicative of flippase activity B . Flippase activity was inhibited by use of NEM in low surface PS cell lines and surface PS levels were measured by annexin V FITC binding assay, by flow cytometry. The graph shows annexin V FITC fold change compared to mock treated cells. C . High surface PS cells were either treated with NEM or left untreated and incubated with NBD-PS for indicated time periods and subjected to BSA extraction and sodium dithionite treatment. % nonextractable NBD-PS (after BSA extraction and sodium dithionite treatment) represents internalized NBD-PS, indicative of flippase activity D . Flippase activity was inhibited by use of NEM in the high surface PS cell lines and surface PS levels were measured by annexin V FITC binding assay by flow cytometry. The graph shows annexin V FITC fold change compared to mock treated cells.

    Article Snippet: The remaining half was spun down to remove non inserted NBD-PS and subjected to BSA extraction of NBD-PS from the outer leaflet by adding 3% fatty acid free BSA (MP biomedicals) in flippase assay buffer.

    Techniques: Inhibition, Activity Assay, Incubation, Binding Assay, Flow Cytometry, Cytometry

    The effect of VASP on lipid metabolism in AML12 cells. A : AML12 cells were transduced with VASP (VASP-OE) or control (ctl) (empty) vector. B : RT-PCR analysis of lipid metabolism–related genes in AML12 cells. Expression of Gapdh as shown by the ratio of CT value ( n = 3). C : Rate of [1- 14 C]palmitate incorporation into acid-soluble metabolites ( n = 4). D : Oleic acid (either 0.1 or 0.4 mmol/L)-induced accumulation of TG in AML12 cells. BSA (1.3 mmol/L) was used as a control ( n = 3). * P

    Journal: Diabetes

    Article Title: VASP Increases Hepatic Fatty Acid Oxidation by Activating AMPK in Mice

    doi: 10.2337/db12-0325

    Figure Lengend Snippet: The effect of VASP on lipid metabolism in AML12 cells. A : AML12 cells were transduced with VASP (VASP-OE) or control (ctl) (empty) vector. B : RT-PCR analysis of lipid metabolism–related genes in AML12 cells. Expression of Gapdh as shown by the ratio of CT value ( n = 3). C : Rate of [1- 14 C]palmitate incorporation into acid-soluble metabolites ( n = 4). D : Oleic acid (either 0.1 or 0.4 mmol/L)-induced accumulation of TG in AML12 cells. BSA (1.3 mmol/L) was used as a control ( n = 3). * P

    Article Snippet: Briefly, AML12 cells were incubated in Dulbecco’s modified Eagle’s medium plus 0.5% fatty acid–free BSA with 0.05 mmol/L carnitine and 0.25 μCi [1-14 C] palmitate (GE Healthcare Life Sciences; Pittsburgh, PA) per well in six-well plates.

    Techniques: Transduction, CTL Assay, Plasmid Preparation, Reverse Transcription Polymerase Chain Reaction, Expressing

    Glutathionation in A549 cells supplemented with BSA-conjugated fatty acids (LA, DHA, BSA carrier control). Cells were treated with BSO or GSH in combination with each of the fatty acids for 48 µM BaP for 24 h. Data represents mean normalized intensity ± SEM of at least 30 images per treatment and 30 cells per image. * indicates significant difference from the corresponding control using Tukey test at p

    Journal: PLoS ONE

    Article Title: Effects of Fatty Acids on Benzo[a]pyrene Uptake and Metabolism in Human Lung Adenocarcinoma A549 Cells

    doi: 10.1371/journal.pone.0090908

    Figure Lengend Snippet: Glutathionation in A549 cells supplemented with BSA-conjugated fatty acids (LA, DHA, BSA carrier control). Cells were treated with BSO or GSH in combination with each of the fatty acids for 48 µM BaP for 24 h. Data represents mean normalized intensity ± SEM of at least 30 images per treatment and 30 cells per image. * indicates significant difference from the corresponding control using Tukey test at p

    Article Snippet: Materials Dulbecco's Modified Eagle's Medium/Ham's Nutrient Mixture F-12 (DMEM-F12), fatty acid free bovine serum albumin (BSA), Dulbecco's phosphate buffered saline (PBS), glutathione (GSH), benzo[a]pyrene (BaP), L- buthionine sulfoximine (BSO), janus green, β-glucuronidase (keyhole limpet, #G8132), triclosan (Irgasan), resorufin, ethyl ether, 3,3′-methylene-bis(4-hydroxycoumarin) (dicumarol), and trans-3,4′,-5-trihydroxystilebene; 3,4′,5-stilbenetriol (resveratrol) were purchased from Sigma-Aldrich Chemical Co. (St. Louis, MO).

    Techniques:

    Effect of 24 hour exposure to fatty acids on human BMMSC energy metabolism. A) Palmitate oxidation, B) palmitate uptake, C) glycolysis, and D) glucose oxidation were measured in human BMMSCs that had been treated for 24 hr with either 0.55 mM albumin (BSA group) or 0.55 mM albumin and 0.4 mM palmitate and/or 0.4 mM oleate prior to these metabolism measurements being made. n = 5–8 The graphs indicate which groups were exposed to these different treatments for the 24 hr prior to the metabolism measurements. During each assay all groups were given Krebs buffer supplemented with 5 mM glucose and 0.4 mM palmitate bound to 0.55 mM albumin. In addition, the Krebs buffer was supplemented with either [U- 14 C]glucose, [1– 14 C]palmitate, or [5– 3 H]glucose for the measurement of glucose oxidation, palmitate oxidation and uptake, or glycolysis, respectively. E) The contribution of metabolic pathways to ATP production were calculated from the metabolic rate results. * Significantly different from all groups. Values are shown as mean ± SEM.

    Journal: PLoS ONE

    Article Title: Effect of Fatty Acids on Human Bone Marrow Mesenchymal Stem Cell Energy Metabolism and Survival

    doi: 10.1371/journal.pone.0120257

    Figure Lengend Snippet: Effect of 24 hour exposure to fatty acids on human BMMSC energy metabolism. A) Palmitate oxidation, B) palmitate uptake, C) glycolysis, and D) glucose oxidation were measured in human BMMSCs that had been treated for 24 hr with either 0.55 mM albumin (BSA group) or 0.55 mM albumin and 0.4 mM palmitate and/or 0.4 mM oleate prior to these metabolism measurements being made. n = 5–8 The graphs indicate which groups were exposed to these different treatments for the 24 hr prior to the metabolism measurements. During each assay all groups were given Krebs buffer supplemented with 5 mM glucose and 0.4 mM palmitate bound to 0.55 mM albumin. In addition, the Krebs buffer was supplemented with either [U- 14 C]glucose, [1– 14 C]palmitate, or [5– 3 H]glucose for the measurement of glucose oxidation, palmitate oxidation and uptake, or glycolysis, respectively. E) The contribution of metabolic pathways to ATP production were calculated from the metabolic rate results. * Significantly different from all groups. Values are shown as mean ± SEM.

    Article Snippet: During experiments assessing the chronic effect of fatty acids on BMMSCs this media was also supplemented with 4% fatty acid free bovine serum albumin (BSA) (Equitech-Bio Inc BAH66) or 4% BSA bound to the indicated type and concentration of fatty acid (Palmitate, Sigma P9767; Oleate, Fluka Analytical 60420; Stearate, Sigma S3381).

    Techniques:

    Effect of acute exposure to fatty acids on human BMMSC energy metabolism. A) Glycolysis, B) glucose oxidation, C) palmitate oxidation, D) palmitate uptake, E) oleate oxidation, and F) oleate uptake were measured in untreated human bone marrow mesenchymal stem cells (BMMSCs). n = 5–7 During each assay Krebs Henseleit buffer was supplemented with 5 mM glucose and, as indicated in each graph, either 0.55 mM albumin (BSA group) or 0.55 mM albumin bound to 0.4 mM palmitate and/or 0.4 mM oleate. In addition, the Krebs Henseleit buffer was supplemented with either [U- 14 C]glucose, [1– 14 C]palmitate, [1– 14 C]oleate, or [5– 3 H]glucose for the measurement of glucose oxidation, palmitate oxidation and uptake, oleate oxidation and uptake, or glycolysis, respectively. Since these experiments assessed the acute effect of palmitate and oleate, BMMSCs were maintained in cell culture media used to culture these immediately up to the start of each assay when the media was switched to Krebs Henseleit buffer supplemented with fatty acids. The levels and type of fatty acid BMMSCs were exposed to is indicated on the x-axis of the figures. * Significantly different from all groups. Values are shown as the mean ± SEM.

    Journal: PLoS ONE

    Article Title: Effect of Fatty Acids on Human Bone Marrow Mesenchymal Stem Cell Energy Metabolism and Survival

    doi: 10.1371/journal.pone.0120257

    Figure Lengend Snippet: Effect of acute exposure to fatty acids on human BMMSC energy metabolism. A) Glycolysis, B) glucose oxidation, C) palmitate oxidation, D) palmitate uptake, E) oleate oxidation, and F) oleate uptake were measured in untreated human bone marrow mesenchymal stem cells (BMMSCs). n = 5–7 During each assay Krebs Henseleit buffer was supplemented with 5 mM glucose and, as indicated in each graph, either 0.55 mM albumin (BSA group) or 0.55 mM albumin bound to 0.4 mM palmitate and/or 0.4 mM oleate. In addition, the Krebs Henseleit buffer was supplemented with either [U- 14 C]glucose, [1– 14 C]palmitate, [1– 14 C]oleate, or [5– 3 H]glucose for the measurement of glucose oxidation, palmitate oxidation and uptake, oleate oxidation and uptake, or glycolysis, respectively. Since these experiments assessed the acute effect of palmitate and oleate, BMMSCs were maintained in cell culture media used to culture these immediately up to the start of each assay when the media was switched to Krebs Henseleit buffer supplemented with fatty acids. The levels and type of fatty acid BMMSCs were exposed to is indicated on the x-axis of the figures. * Significantly different from all groups. Values are shown as the mean ± SEM.

    Article Snippet: During experiments assessing the chronic effect of fatty acids on BMMSCs this media was also supplemented with 4% fatty acid free bovine serum albumin (BSA) (Equitech-Bio Inc BAH66) or 4% BSA bound to the indicated type and concentration of fatty acid (Palmitate, Sigma P9767; Oleate, Fluka Analytical 60420; Stearate, Sigma S3381).

    Techniques: Cell Culture

    Effect of 24 hr exposure to palmitate and oleate on human BMMSC mitochondrial membrane potential. A) Images of tetramethylrhodamine methyl ester (TMRM) stained bone marrow mesenchymal stem cells (BMMSCs) treated for 24 hr with indicated treatments. B) Relative TMRM levels. BMMSCs were treated for 24 hr with 0.55 mM bovine serum albumin (BSA) alone or palmitate and/or oleate bound to 0.55 mM albumin. TMRM and Hoechst stain were added to the medium to measure mitochondrial membrane potential and stain nuclei, respectively, and images were taken. All fatty acid treated groups were also treated with media supplemented with 0.55 mM albumin in addition to the type and amount of fatty acid indicated in the figures. 4 separate observations. Values are shown as the mean ± SEM. * Significantly different from the BSA group.

    Journal: PLoS ONE

    Article Title: Effect of Fatty Acids on Human Bone Marrow Mesenchymal Stem Cell Energy Metabolism and Survival

    doi: 10.1371/journal.pone.0120257

    Figure Lengend Snippet: Effect of 24 hr exposure to palmitate and oleate on human BMMSC mitochondrial membrane potential. A) Images of tetramethylrhodamine methyl ester (TMRM) stained bone marrow mesenchymal stem cells (BMMSCs) treated for 24 hr with indicated treatments. B) Relative TMRM levels. BMMSCs were treated for 24 hr with 0.55 mM bovine serum albumin (BSA) alone or palmitate and/or oleate bound to 0.55 mM albumin. TMRM and Hoechst stain were added to the medium to measure mitochondrial membrane potential and stain nuclei, respectively, and images were taken. All fatty acid treated groups were also treated with media supplemented with 0.55 mM albumin in addition to the type and amount of fatty acid indicated in the figures. 4 separate observations. Values are shown as the mean ± SEM. * Significantly different from the BSA group.

    Article Snippet: During experiments assessing the chronic effect of fatty acids on BMMSCs this media was also supplemented with 4% fatty acid free bovine serum albumin (BSA) (Equitech-Bio Inc BAH66) or 4% BSA bound to the indicated type and concentration of fatty acid (Palmitate, Sigma P9767; Oleate, Fluka Analytical 60420; Stearate, Sigma S3381).

    Techniques: Staining