anti-mouse flow cytometry antibodies Search Results


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  • 99
    Thermo Fisher flow cytometry mice
    HSCs from sleep-deprived mice have reduced homing capacity in vivo and in vitro (a) HSCs (2000 FACS-purified cells) derived from GFP-expressing, sleep-deprived mice were transplanted into lethally irradiated congenic mice that did not express GFP (control mice were allowed to sleep for the same duration). The donor HSCs were visualized 12 hours later in the recipient’s bone, under a fluorescence microscope. GFP expression was validated using anti-GFP staining and a representative image is shown in the insert. Scale 10µm (n=6–11 mice per group; mean±s.e.m). ( b ) The number of GFP-labeled cells in the recipient’s bone marrow was determined. Results are presented as the percentage of homing, assuming that two tibias and two femurs represent 20% of total mouse bone marrow (student’s t -test ; p=0.021; t=2.56; df=15 ). ( c ) Migration towards SDF-1α was determined in vitro using a transwell migration assay. KLS cells were placed in the upper chamber and the chemoattractant (SDF-1α, 50, 100, or 200ng/mL) was immersed in the medium of the lower chamber. Migration across the membrane in response to the chemoattractant was determined using flow <t>cytometry,</t> after four hours of incubation, and compared to baseline migration in the absence of the chemoattractant. Results are presented as a migration index (50ng/mL SDF-1α: 9.6 ± 1.7% in the sleep group compared to 2.06 ± 0.4% in the sleep-deprived group; 100ng/mL SDF-1α: 14.5 ± 3.4% in the sleep compared to 3.6 ± 1.4% in the sleep-deprived mice; Repeated measures ANOVA- sleep: F (2,15)=10.18, p
    Flow Cytometry Mice, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 29 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Millipore anti mouse tlr4 n term antibody
    Dynamin activity mediates <t>TLR4-dependent</t> ERK 1/2 phosphorylation, c - fos and TNF mRNA accumulation and TNF secretion. WT BMMCs were pre-incubated with dynasore (80 μM) for 30 min before LPS addition (500 ng/mL). After stimulus, the cells were incubated at 37 °C for the indicated times and processed to obtain total protein or total RNA for RT-PCR. a ERK1/2 phosphorylation, b c - fos mRNA accumulation, c TNF mRNA synthesis, and d TNF secretion. Data are presented as the mean ± SD of 3–4 independent experiments. p
    Anti Mouse Tlr4 N Term Antibody, supplied by Millipore, used in various techniques. Bioz Stars score: 94/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    Becton Dickinson flow cytometry anti mouse cd3
    T cell recovery after irradiation-induced lymphopenia. B6 mice, Fas fl/fl lck-cre, and Fas fl/fl mice were irradiated with 6 Gy, <t>CD3</t> T cells in the spleen were counted and analyzed by flow <t>cytometry.</t> Approximately 3–4 mice per group per time point (5-time points total) were analyzed.
    Flow Cytometry Anti Mouse Cd3, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 85/100, based on 19 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Becton Dickinson anti mouse flow cytometry antibodies
    Ectopic Sestrin2 expression in M1 macrophages attenuates post-MI inflammation and favors tissue repair in hearts. (A) Macrophage abundance in infarcted myocardium at day 3 after myocardial infarction (MI) (day 4 after Clophosome ® treatment). Ctrl: un-pretreated mice. Depletion: Clophosome ® injection. Depletion + M (L-C): Clophosome ® injection followed by transfer of M1 macrophages transduced with control lentivirus. Depletion + M (L-S): Clophosome ® injection followed by transfer of M1 macrophages transduced with SESN2 lentiviral activation particles. Numbers in the plots are proportions of gated cells populations. This is a representative of two independent experiments. (B,C) mRNA abundance of indicated cytokines and chemokines in infarcted myocardium. (D,E) Percentage of Gr-1 + neutrophils in the whole cardiac cells at day 3 after MI. Representative flow <t>cytometry</t> dot plots are shown in (D) , and statistics is shown in (E) . (F) Masson staining at day 7 after MI. Left panel: representative images. Right panel: statistics of collagen volume fraction (CVF). N = 4–5 per group. * p
    Anti Mouse Flow Cytometry Antibodies, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 92/100, based on 26 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Thermo Fisher anti mouse ki 67 pe flow cytometric antibody
    LINC00982 and LL22NC03-N14H11.1 have opposite roles in the proliferation of gastric cancer cells. (A) Reverse transcription-quantitative polymerase chain reaction revealed that the expression of LINC00982 in the LINC00982-overexpressed SGC-7901 cell line (left) and the expression of LL22NC03-N14H11.1 in the LL22NC03-N14H11.1-down-regulated SGC-7901 cell line (right). (B) The MTT assay revealed the proliferative status of the SGC-7901 cell line after LINC00982 being overexpressed or LL22NC03-N14H11.1 being downregulated. (C) Flow <t>cytometric</t> analysis of the expression of <t>Ki-67</t> in the SGC-7901 cell line after LINC00982 was overexpressed or LL22NC03-N14H11.1 was downregulated. All the experiments were replicated a minimum of 3. The error bars represent the standard deviation. *P
    Anti Mouse Ki 67 Pe Flow Cytometric Antibody, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 92/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    96
    R&D Systems flow cytometry mouse anti human axl mab
    ZIKV infection of unstimulated and decidualized T-HESC cell line. (A) T-HESC (left) and decidualized (d)T-HESC (right) were stained with anti-dsRNA or Flavi E mAbs whereas the nuclei were stained with Hoechst. Scale bar: 20 µm. (B) Surface expression of <t>AXL</t> (red) and MER (blue) in T-HESC (left) and dT-HESC (right) was determined by flow <t>cytometry.</t> The histograms of one experiment representative of 3 independently performed are shown. Double immunostaining for Flavi E and calreticulin (C) or vimentin (D) in T-HESC (left) and dT-HESC (right) either uninfected or infected with MR766 at 72 h post-infection; Hoechst was used to stain nuclei. Scale bar: 10 µm.
    Flow Cytometry Mouse Anti Human Axl Mab, supplied by R&D Systems, used in various techniques. Bioz Stars score: 96/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    BioLegend pe anti mouse cd150 slam
    ZIKV infection of unstimulated and decidualized T-HESC cell line. (A) T-HESC (left) and decidualized (d)T-HESC (right) were stained with anti-dsRNA or Flavi E mAbs whereas the nuclei were stained with Hoechst. Scale bar: 20 µm. (B) Surface expression of <t>AXL</t> (red) and MER (blue) in T-HESC (left) and dT-HESC (right) was determined by flow <t>cytometry.</t> The histograms of one experiment representative of 3 independently performed are shown. Double immunostaining for Flavi E and calreticulin (C) or vimentin (D) in T-HESC (left) and dT-HESC (right) either uninfected or infected with MR766 at 72 h post-infection; Hoechst was used to stain nuclei. Scale bar: 10 µm.
    Pe Anti Mouse Cd150 Slam, supplied by BioLegend, used in various techniques. Bioz Stars score: 99/100, based on 160 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    95
    BioLegend pe anti mouse ly 6c
    ZIKV infection of unstimulated and decidualized T-HESC cell line. (A) T-HESC (left) and decidualized (d)T-HESC (right) were stained with anti-dsRNA or Flavi E mAbs whereas the nuclei were stained with Hoechst. Scale bar: 20 µm. (B) Surface expression of <t>AXL</t> (red) and MER (blue) in T-HESC (left) and dT-HESC (right) was determined by flow <t>cytometry.</t> The histograms of one experiment representative of 3 independently performed are shown. Double immunostaining for Flavi E and calreticulin (C) or vimentin (D) in T-HESC (left) and dT-HESC (right) either uninfected or infected with MR766 at 72 h post-infection; Hoechst was used to stain nuclei. Scale bar: 10 µm.
    Pe Anti Mouse Ly 6c, supplied by BioLegend, used in various techniques. Bioz Stars score: 95/100, based on 36 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    97
    BioLegend alexa fluor 647 anti mouse cd3
    ZIKV infection of unstimulated and decidualized T-HESC cell line. (A) T-HESC (left) and decidualized (d)T-HESC (right) were stained with anti-dsRNA or Flavi E mAbs whereas the nuclei were stained with Hoechst. Scale bar: 20 µm. (B) Surface expression of <t>AXL</t> (red) and MER (blue) in T-HESC (left) and dT-HESC (right) was determined by flow <t>cytometry.</t> The histograms of one experiment representative of 3 independently performed are shown. Double immunostaining for Flavi E and calreticulin (C) or vimentin (D) in T-HESC (left) and dT-HESC (right) either uninfected or infected with MR766 at 72 h post-infection; Hoechst was used to stain nuclei. Scale bar: 10 µm.
    Alexa Fluor 647 Anti Mouse Cd3, supplied by BioLegend, used in various techniques. Bioz Stars score: 97/100, based on 38 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Thermo Fisher anti human cytokeratine 18 fitc prelabeled monoclonal mouse flow cytometry antibody
    ZIKV infection of unstimulated and decidualized T-HESC cell line. (A) T-HESC (left) and decidualized (d)T-HESC (right) were stained with anti-dsRNA or Flavi E mAbs whereas the nuclei were stained with Hoechst. Scale bar: 20 µm. (B) Surface expression of <t>AXL</t> (red) and MER (blue) in T-HESC (left) and dT-HESC (right) was determined by flow <t>cytometry.</t> The histograms of one experiment representative of 3 independently performed are shown. Double immunostaining for Flavi E and calreticulin (C) or vimentin (D) in T-HESC (left) and dT-HESC (right) either uninfected or infected with MR766 at 72 h post-infection; Hoechst was used to stain nuclei. Scale bar: 10 µm.
    Anti Human Cytokeratine 18 Fitc Prelabeled Monoclonal Mouse Flow Cytometry Antibody, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 93/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    96
    Millipore anti mouse igg fab specific f ab 2 fragment fitc antibody produced in goat
    ZIKV infection of unstimulated and decidualized T-HESC cell line. (A) T-HESC (left) and decidualized (d)T-HESC (right) were stained with anti-dsRNA or Flavi E mAbs whereas the nuclei were stained with Hoechst. Scale bar: 20 µm. (B) Surface expression of <t>AXL</t> (red) and MER (blue) in T-HESC (left) and dT-HESC (right) was determined by flow <t>cytometry.</t> The histograms of one experiment representative of 3 independently performed are shown. Double immunostaining for Flavi E and calreticulin (C) or vimentin (D) in T-HESC (left) and dT-HESC (right) either uninfected or infected with MR766 at 72 h post-infection; Hoechst was used to stain nuclei. Scale bar: 10 µm.
    Anti Mouse Igg Fab Specific F Ab 2 Fragment Fitc Antibody Produced In Goat, supplied by Millipore, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    95
    Cedarlane anti rat cd62l fitc clone ox 85 mouse igg1
    ZIKV infection of unstimulated and decidualized T-HESC cell line. (A) T-HESC (left) and decidualized (d)T-HESC (right) were stained with anti-dsRNA or Flavi E mAbs whereas the nuclei were stained with Hoechst. Scale bar: 20 µm. (B) Surface expression of <t>AXL</t> (red) and MER (blue) in T-HESC (left) and dT-HESC (right) was determined by flow <t>cytometry.</t> The histograms of one experiment representative of 3 independently performed are shown. Double immunostaining for Flavi E and calreticulin (C) or vimentin (D) in T-HESC (left) and dT-HESC (right) either uninfected or infected with MR766 at 72 h post-infection; Hoechst was used to stain nuclei. Scale bar: 10 µm.
    Anti Rat Cd62l Fitc Clone Ox 85 Mouse Igg1, supplied by Cedarlane, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore monoclonal anti actin alpha smooth muscle fitc antibody
    ZIKV infection of unstimulated and decidualized T-HESC cell line. (A) T-HESC (left) and decidualized (d)T-HESC (right) were stained with anti-dsRNA or Flavi E mAbs whereas the nuclei were stained with Hoechst. Scale bar: 20 µm. (B) Surface expression of <t>AXL</t> (red) and MER (blue) in T-HESC (left) and dT-HESC (right) was determined by flow <t>cytometry.</t> The histograms of one experiment representative of 3 independently performed are shown. Double immunostaining for Flavi E and calreticulin (C) or vimentin (D) in T-HESC (left) and dT-HESC (right) either uninfected or infected with MR766 at 72 h post-infection; Hoechst was used to stain nuclei. Scale bar: 10 µm.
    Monoclonal Anti Actin Alpha Smooth Muscle Fitc Antibody, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 1230 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    97
    BioLegend percp cy5 5 anti mouse human cd45r b220
    ZIKV infection of unstimulated and decidualized T-HESC cell line. (A) T-HESC (left) and decidualized (d)T-HESC (right) were stained with anti-dsRNA or Flavi E mAbs whereas the nuclei were stained with Hoechst. Scale bar: 20 µm. (B) Surface expression of <t>AXL</t> (red) and MER (blue) in T-HESC (left) and dT-HESC (right) was determined by flow <t>cytometry.</t> The histograms of one experiment representative of 3 independently performed are shown. Double immunostaining for Flavi E and calreticulin (C) or vimentin (D) in T-HESC (left) and dT-HESC (right) either uninfected or infected with MR766 at 72 h post-infection; Hoechst was used to stain nuclei. Scale bar: 10 µm.
    Percp Cy5 5 Anti Mouse Human Cd45r B220, supplied by BioLegend, used in various techniques. Bioz Stars score: 97/100, based on 48 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    90
    Thermo Fisher flow cytometric analysis anti human mouse cd44 fitc
    ZIKV infection of unstimulated and decidualized T-HESC cell line. (A) T-HESC (left) and decidualized (d)T-HESC (right) were stained with anti-dsRNA or Flavi E mAbs whereas the nuclei were stained with Hoechst. Scale bar: 20 µm. (B) Surface expression of <t>AXL</t> (red) and MER (blue) in T-HESC (left) and dT-HESC (right) was determined by flow <t>cytometry.</t> The histograms of one experiment representative of 3 independently performed are shown. Double immunostaining for Flavi E and calreticulin (C) or vimentin (D) in T-HESC (left) and dT-HESC (right) either uninfected or infected with MR766 at 72 h post-infection; Hoechst was used to stain nuclei. Scale bar: 10 µm.
    Flow Cytometric Analysis Anti Human Mouse Cd44 Fitc, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 90/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    90
    Becton Dickinson flow cytometric analysis pe conjugated mouse anti human ox40l
    ZIKV infection of unstimulated and decidualized T-HESC cell line. (A) T-HESC (left) and decidualized (d)T-HESC (right) were stained with anti-dsRNA or Flavi E mAbs whereas the nuclei were stained with Hoechst. Scale bar: 20 µm. (B) Surface expression of <t>AXL</t> (red) and MER (blue) in T-HESC (left) and dT-HESC (right) was determined by flow <t>cytometry.</t> The histograms of one experiment representative of 3 independently performed are shown. Double immunostaining for Flavi E and calreticulin (C) or vimentin (D) in T-HESC (left) and dT-HESC (right) either uninfected or infected with MR766 at 72 h post-infection; Hoechst was used to stain nuclei. Scale bar: 10 µm.
    Flow Cytometric Analysis Pe Conjugated Mouse Anti Human Ox40l, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 90/100, based on 22 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    R&D Systems allophycocyanin conjugated anti mouse cd151 flow cytometry ab
    <t>CD151</t> deficiency does not alter basal phenotype of cultured BMMCs. ( A ) CD151 deficiency does not affect growth dynamics of mast cell culture. WT and CD151 −/− BMMCs were cultured in IL-3–conditioned media for 8 wk and the total numbers of live cells in culture were counted at each time point indicated ( left ). Flow <t>cytometry</t> for CD151 expression on peritoneal mast cells (flow cytometry chart) ( middle ), qPCR detection of CD151 mRNA in WT and CD151 −/− BMMCs (bar graph, right ), and Western blot analysis of total-protein lysates for CD151 expression (Western blot, right ) all confirm constitutive CD151 expression in WT mast cells. WT peritoneal mast cells are shown as gray-filled histogram with dotted line and negative control as transparent histogram with dotted line. In immunoblotting, actin was used as a loading control. ( B ) Five-week-old BMMCs from WT and CD151 −/− mice were stained with toluidine blue and images were obtained with an original magnification of ×100. Flow cytometry analysis of FcεRI and c-Kit surface expression and purity of WT and CD151 −/− BMMC cultures. All data are representative of three independent experiments. Data are represented as mean ± SEM. * p
    Allophycocyanin Conjugated Anti Mouse Cd151 Flow Cytometry Ab, supplied by R&D Systems, used in various techniques. Bioz Stars score: 88/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Thermo Fisher click it plus edu alexa fluor 647 flow cytometry assay kit
    <t>CD151</t> deficiency does not alter basal phenotype of cultured BMMCs. ( A ) CD151 deficiency does not affect growth dynamics of mast cell culture. WT and CD151 −/− BMMCs were cultured in IL-3–conditioned media for 8 wk and the total numbers of live cells in culture were counted at each time point indicated ( left ). Flow <t>cytometry</t> for CD151 expression on peritoneal mast cells (flow cytometry chart) ( middle ), qPCR detection of CD151 mRNA in WT and CD151 −/− BMMCs (bar graph, right ), and Western blot analysis of total-protein lysates for CD151 expression (Western blot, right ) all confirm constitutive CD151 expression in WT mast cells. WT peritoneal mast cells are shown as gray-filled histogram with dotted line and negative control as transparent histogram with dotted line. In immunoblotting, actin was used as a loading control. ( B ) Five-week-old BMMCs from WT and CD151 −/− mice were stained with toluidine blue and images were obtained with an original magnification of ×100. Flow cytometry analysis of FcεRI and c-Kit surface expression and purity of WT and CD151 −/− BMMC cultures. All data are representative of three independent experiments. Data are represented as mean ± SEM. * p
    Click It Plus Edu Alexa Fluor 647 Flow Cytometry Assay Kit, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 615 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    84
    Becton Dickinson flow cytometry a pe conjugated mouse anti human cd184 cxcr4 antibody
    <t>CD151</t> deficiency does not alter basal phenotype of cultured BMMCs. ( A ) CD151 deficiency does not affect growth dynamics of mast cell culture. WT and CD151 −/− BMMCs were cultured in IL-3–conditioned media for 8 wk and the total numbers of live cells in culture were counted at each time point indicated ( left ). Flow <t>cytometry</t> for CD151 expression on peritoneal mast cells (flow cytometry chart) ( middle ), qPCR detection of CD151 mRNA in WT and CD151 −/− BMMCs (bar graph, right ), and Western blot analysis of total-protein lysates for CD151 expression (Western blot, right ) all confirm constitutive CD151 expression in WT mast cells. WT peritoneal mast cells are shown as gray-filled histogram with dotted line and negative control as transparent histogram with dotted line. In immunoblotting, actin was used as a loading control. ( B ) Five-week-old BMMCs from WT and CD151 −/− mice were stained with toluidine blue and images were obtained with an original magnification of ×100. Flow cytometry analysis of FcεRI and c-Kit surface expression and purity of WT and CD151 −/− BMMC cultures. All data are representative of three independent experiments. Data are represented as mean ± SEM. * p
    Flow Cytometry A Pe Conjugated Mouse Anti Human Cd184 Cxcr4 Antibody, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 84/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    Thermo Fisher flow cytometry alexa fluor 647 conjugated armenian hamster anti mouse cd11c
    <t>CD151</t> deficiency does not alter basal phenotype of cultured BMMCs. ( A ) CD151 deficiency does not affect growth dynamics of mast cell culture. WT and CD151 −/− BMMCs were cultured in IL-3–conditioned media for 8 wk and the total numbers of live cells in culture were counted at each time point indicated ( left ). Flow <t>cytometry</t> for CD151 expression on peritoneal mast cells (flow cytometry chart) ( middle ), qPCR detection of CD151 mRNA in WT and CD151 −/− BMMCs (bar graph, right ), and Western blot analysis of total-protein lysates for CD151 expression (Western blot, right ) all confirm constitutive CD151 expression in WT mast cells. WT peritoneal mast cells are shown as gray-filled histogram with dotted line and negative control as transparent histogram with dotted line. In immunoblotting, actin was used as a loading control. ( B ) Five-week-old BMMCs from WT and CD151 −/− mice were stained with toluidine blue and images were obtained with an original magnification of ×100. Flow cytometry analysis of FcεRI and c-Kit surface expression and purity of WT and CD151 −/− BMMC cultures. All data are representative of three independent experiments. Data are represented as mean ± SEM. * p
    Flow Cytometry Alexa Fluor 647 Conjugated Armenian Hamster Anti Mouse Cd11c, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 85/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    BioLegend flow cytometry analysis
    Extended validation of candidate genes displaying stronger dropout effects in STAG2 -null versus wild-type cells. (A) Cas9-GFP–expressing KBM-7 clonal cell line (B4) was lentivirally transduced with a validated sgRNA targeting STAG2 , and several subclones were isolated and characterized for STAG2 knockout by immunoblotting. (B) For each candidate gene, multiple sgRNAs scoring in the primary screen (2 or 3, as indicated) were tested in parallel competitive proliferation assays in STAG2 –wild-type (B4, blue lines) and STAG2 -mutant (c9, red lines) cell lines used in the screen and in an independent STAG2 -mutant clone (c11, orange lines). Cells were transduced with lentiviral vectors co-expressing mCherry and an sgRNA targeting the indicated genes, and the fraction of mCherry+ cells was monitored over time using flow <t>cytometry.</t> Plotted are averages of 2–3 independent sgRNAs analyzed for each gene; error bars represent standard deviations. Note that knockout of most candidates (e.g., BUB1, PAXIP1, PDCD5, PP2R1A, and SKA3) was deleterious in both STAG2–wild-type and STAG2-mutant cells, with some differences in depletion dynamics. In other instances, knockout effects turned out to be specific without being strongly detrimental in STAG2-mutant cells (e.g., CHMP7, FLI1, and SKI), and in some cases, they could not be confirmed in an independent STAG2-null clone (c11, e.g., DNAJC9 and FANCI), indicating that clonal artifacts remain an issue even when starting out from a single-cell–derived clone.
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    Becton Dickinson flow cytometry apc mouse anti cd184
    Extended validation of candidate genes displaying stronger dropout effects in STAG2 -null versus wild-type cells. (A) Cas9-GFP–expressing KBM-7 clonal cell line (B4) was lentivirally transduced with a validated sgRNA targeting STAG2 , and several subclones were isolated and characterized for STAG2 knockout by immunoblotting. (B) For each candidate gene, multiple sgRNAs scoring in the primary screen (2 or 3, as indicated) were tested in parallel competitive proliferation assays in STAG2 –wild-type (B4, blue lines) and STAG2 -mutant (c9, red lines) cell lines used in the screen and in an independent STAG2 -mutant clone (c11, orange lines). Cells were transduced with lentiviral vectors co-expressing mCherry and an sgRNA targeting the indicated genes, and the fraction of mCherry+ cells was monitored over time using flow <t>cytometry.</t> Plotted are averages of 2–3 independent sgRNAs analyzed for each gene; error bars represent standard deviations. Note that knockout of most candidates (e.g., BUB1, PAXIP1, PDCD5, PP2R1A, and SKA3) was deleterious in both STAG2–wild-type and STAG2-mutant cells, with some differences in depletion dynamics. In other instances, knockout effects turned out to be specific without being strongly detrimental in STAG2-mutant cells (e.g., CHMP7, FLI1, and SKI), and in some cases, they could not be confirmed in an independent STAG2-null clone (c11, e.g., DNAJC9 and FANCI), indicating that clonal artifacts remain an issue even when starting out from a single-cell–derived clone.
    Flow Cytometry Apc Mouse Anti Cd184, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 91/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Becton Dickinson flow cytometry anti mouse cd45
    Extended validation of candidate genes displaying stronger dropout effects in STAG2 -null versus wild-type cells. (A) Cas9-GFP–expressing KBM-7 clonal cell line (B4) was lentivirally transduced with a validated sgRNA targeting STAG2 , and several subclones were isolated and characterized for STAG2 knockout by immunoblotting. (B) For each candidate gene, multiple sgRNAs scoring in the primary screen (2 or 3, as indicated) were tested in parallel competitive proliferation assays in STAG2 –wild-type (B4, blue lines) and STAG2 -mutant (c9, red lines) cell lines used in the screen and in an independent STAG2 -mutant clone (c11, orange lines). Cells were transduced with lentiviral vectors co-expressing mCherry and an sgRNA targeting the indicated genes, and the fraction of mCherry+ cells was monitored over time using flow <t>cytometry.</t> Plotted are averages of 2–3 independent sgRNAs analyzed for each gene; error bars represent standard deviations. Note that knockout of most candidates (e.g., BUB1, PAXIP1, PDCD5, PP2R1A, and SKA3) was deleterious in both STAG2–wild-type and STAG2-mutant cells, with some differences in depletion dynamics. In other instances, knockout effects turned out to be specific without being strongly detrimental in STAG2-mutant cells (e.g., CHMP7, FLI1, and SKI), and in some cases, they could not be confirmed in an independent STAG2-null clone (c11, e.g., DNAJC9 and FANCI), indicating that clonal artifacts remain an issue even when starting out from a single-cell–derived clone.
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    Thermo Fisher flow cytometry anti human mouse phospho src y418 percp efluor 710
    Extended validation of candidate genes displaying stronger dropout effects in STAG2 -null versus wild-type cells. (A) Cas9-GFP–expressing KBM-7 clonal cell line (B4) was lentivirally transduced with a validated sgRNA targeting STAG2 , and several subclones were isolated and characterized for STAG2 knockout by immunoblotting. (B) For each candidate gene, multiple sgRNAs scoring in the primary screen (2 or 3, as indicated) were tested in parallel competitive proliferation assays in STAG2 –wild-type (B4, blue lines) and STAG2 -mutant (c9, red lines) cell lines used in the screen and in an independent STAG2 -mutant clone (c11, orange lines). Cells were transduced with lentiviral vectors co-expressing mCherry and an sgRNA targeting the indicated genes, and the fraction of mCherry+ cells was monitored over time using flow <t>cytometry.</t> Plotted are averages of 2–3 independent sgRNAs analyzed for each gene; error bars represent standard deviations. Note that knockout of most candidates (e.g., BUB1, PAXIP1, PDCD5, PP2R1A, and SKA3) was deleterious in both STAG2–wild-type and STAG2-mutant cells, with some differences in depletion dynamics. In other instances, knockout effects turned out to be specific without being strongly detrimental in STAG2-mutant cells (e.g., CHMP7, FLI1, and SKI), and in some cases, they could not be confirmed in an independent STAG2-null clone (c11, e.g., DNAJC9 and FANCI), indicating that clonal artifacts remain an issue even when starting out from a single-cell–derived clone.
    Flow Cytometry Anti Human Mouse Phospho Src Y418 Percp Efluor 710, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 84/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    HSCs from sleep-deprived mice have reduced homing capacity in vivo and in vitro (a) HSCs (2000 FACS-purified cells) derived from GFP-expressing, sleep-deprived mice were transplanted into lethally irradiated congenic mice that did not express GFP (control mice were allowed to sleep for the same duration). The donor HSCs were visualized 12 hours later in the recipient’s bone, under a fluorescence microscope. GFP expression was validated using anti-GFP staining and a representative image is shown in the insert. Scale 10µm (n=6–11 mice per group; mean±s.e.m). ( b ) The number of GFP-labeled cells in the recipient’s bone marrow was determined. Results are presented as the percentage of homing, assuming that two tibias and two femurs represent 20% of total mouse bone marrow (student’s t -test ; p=0.021; t=2.56; df=15 ). ( c ) Migration towards SDF-1α was determined in vitro using a transwell migration assay. KLS cells were placed in the upper chamber and the chemoattractant (SDF-1α, 50, 100, or 200ng/mL) was immersed in the medium of the lower chamber. Migration across the membrane in response to the chemoattractant was determined using flow cytometry, after four hours of incubation, and compared to baseline migration in the absence of the chemoattractant. Results are presented as a migration index (50ng/mL SDF-1α: 9.6 ± 1.7% in the sleep group compared to 2.06 ± 0.4% in the sleep-deprived group; 100ng/mL SDF-1α: 14.5 ± 3.4% in the sleep compared to 3.6 ± 1.4% in the sleep-deprived mice; Repeated measures ANOVA- sleep: F (2,15)=10.18, p

    Journal: Nature communications

    Article Title: Sleep disruption impairs hematopoietic stem cell transplantation in mice

    doi: 10.1038/ncomms9516

    Figure Lengend Snippet: HSCs from sleep-deprived mice have reduced homing capacity in vivo and in vitro (a) HSCs (2000 FACS-purified cells) derived from GFP-expressing, sleep-deprived mice were transplanted into lethally irradiated congenic mice that did not express GFP (control mice were allowed to sleep for the same duration). The donor HSCs were visualized 12 hours later in the recipient’s bone, under a fluorescence microscope. GFP expression was validated using anti-GFP staining and a representative image is shown in the insert. Scale 10µm (n=6–11 mice per group; mean±s.e.m). ( b ) The number of GFP-labeled cells in the recipient’s bone marrow was determined. Results are presented as the percentage of homing, assuming that two tibias and two femurs represent 20% of total mouse bone marrow (student’s t -test ; p=0.021; t=2.56; df=15 ). ( c ) Migration towards SDF-1α was determined in vitro using a transwell migration assay. KLS cells were placed in the upper chamber and the chemoattractant (SDF-1α, 50, 100, or 200ng/mL) was immersed in the medium of the lower chamber. Migration across the membrane in response to the chemoattractant was determined using flow cytometry, after four hours of incubation, and compared to baseline migration in the absence of the chemoattractant. Results are presented as a migration index (50ng/mL SDF-1α: 9.6 ± 1.7% in the sleep group compared to 2.06 ± 0.4% in the sleep-deprived group; 100ng/mL SDF-1α: 14.5 ± 3.4% in the sleep compared to 3.6 ± 1.4% in the sleep-deprived mice; Repeated measures ANOVA- sleep: F (2,15)=10.18, p

    Article Snippet: Flow cytometry Mice were euthanized, using CO2, and bone marrow was harvested into PBS containing 2% fetal calf serum (FCS).

    Techniques: Mouse Assay, In Vivo, In Vitro, FACS, Purification, Derivative Assay, Expressing, Irradiation, Fluorescence, Microscopy, Staining, Labeling, Migration, Transwell Migration Assay, Flow Cytometry, Incubation

    HSCs isolated from sleep-deprived mice have reduced mid-term and long-term reconstitution potential in lethally irradiated hosts ( a ) Mice were sleep-deprived, by gentle handling, for four hours, immediately after light onset (ZT0-ZT4; n=8 per group). Rapid eye movement (REM) and non-REM (NREM) duration were determined using electroencephalography (EEG) and electromyography (EMG), individually plotted for each hour (sleep deprived points in red; mean±s.e.m). ( b ) Plasma of the sleep and sleep-deprived mice was analyzed by ELISA for corticosterone levels (n=8 per group; mean±s.e.m). ( c ) We isolated HSCs from mice that were allowed to sleep or sleep-deprived mice. The flow cytometry gating scheme for HSCs isolation is shown on a representative mouse. (d) To test the mid-term and long-term transplantation potential of the isolated HSCs we intravenously injected 300 HSCs mice into lethally irradiated congenic recipients (to distinguish between the donor and recipient cells we used CD45.1 mice as donors and CD45.2 as recipients). Peripheral blood (PB) samples were collected from the recipient mice at ( e ) eight weeks (peripheral blood), ( f ) 16 weeks and ( g ) bone marrow (BM) post-transplantation. Cells were analyzed for myeloid chimerism, as determined by the percentage of myeloid cells derived from the donor compared to the total number of myeloid cells in the indicated tissue (mean±s.e.m; Student’s t -test; *** p

    Journal: Nature communications

    Article Title: Sleep disruption impairs hematopoietic stem cell transplantation in mice

    doi: 10.1038/ncomms9516

    Figure Lengend Snippet: HSCs isolated from sleep-deprived mice have reduced mid-term and long-term reconstitution potential in lethally irradiated hosts ( a ) Mice were sleep-deprived, by gentle handling, for four hours, immediately after light onset (ZT0-ZT4; n=8 per group). Rapid eye movement (REM) and non-REM (NREM) duration were determined using electroencephalography (EEG) and electromyography (EMG), individually plotted for each hour (sleep deprived points in red; mean±s.e.m). ( b ) Plasma of the sleep and sleep-deprived mice was analyzed by ELISA for corticosterone levels (n=8 per group; mean±s.e.m). ( c ) We isolated HSCs from mice that were allowed to sleep or sleep-deprived mice. The flow cytometry gating scheme for HSCs isolation is shown on a representative mouse. (d) To test the mid-term and long-term transplantation potential of the isolated HSCs we intravenously injected 300 HSCs mice into lethally irradiated congenic recipients (to distinguish between the donor and recipient cells we used CD45.1 mice as donors and CD45.2 as recipients). Peripheral blood (PB) samples were collected from the recipient mice at ( e ) eight weeks (peripheral blood), ( f ) 16 weeks and ( g ) bone marrow (BM) post-transplantation. Cells were analyzed for myeloid chimerism, as determined by the percentage of myeloid cells derived from the donor compared to the total number of myeloid cells in the indicated tissue (mean±s.e.m; Student’s t -test; *** p

    Article Snippet: Flow cytometry Mice were euthanized, using CO2, and bone marrow was harvested into PBS containing 2% fetal calf serum (FCS).

    Techniques: Isolation, Mouse Assay, Irradiation, Enzyme-linked Immunosorbent Assay, Flow Cytometry, Transplantation Assay, Injection, Derivative Assay

    Dynamin activity mediates TLR4-dependent ERK 1/2 phosphorylation, c - fos and TNF mRNA accumulation and TNF secretion. WT BMMCs were pre-incubated with dynasore (80 μM) for 30 min before LPS addition (500 ng/mL). After stimulus, the cells were incubated at 37 °C for the indicated times and processed to obtain total protein or total RNA for RT-PCR. a ERK1/2 phosphorylation, b c - fos mRNA accumulation, c TNF mRNA synthesis, and d TNF secretion. Data are presented as the mean ± SD of 3–4 independent experiments. p

    Journal: Journal of Neuroinflammation

    Article Title: Mutant Huntingtin affects toll-like receptor 4 intracellular trafficking and cytokine production in mast cells

    doi: 10.1186/s12974-020-01758-9

    Figure Lengend Snippet: Dynamin activity mediates TLR4-dependent ERK 1/2 phosphorylation, c - fos and TNF mRNA accumulation and TNF secretion. WT BMMCs were pre-incubated with dynasore (80 μM) for 30 min before LPS addition (500 ng/mL). After stimulus, the cells were incubated at 37 °C for the indicated times and processed to obtain total protein or total RNA for RT-PCR. a ERK1/2 phosphorylation, b c - fos mRNA accumulation, c TNF mRNA synthesis, and d TNF secretion. Data are presented as the mean ± SD of 3–4 independent experiments. p

    Article Snippet: Antibodies against TLR-4 receptor used for confocal microscopy were purchased from BioLegend (San Diego, CA, USA) (Cat. No. SA1521, recognizing TLR4/MD2), and those used for flow cytometry (Cat. No. SAB1300056, recognizing N-terminal region of TLR-4 receptor) were purchased from Sigma-Aldrich.

    Techniques: Activity Assay, Incubation, Reverse Transcription Polymerase Chain Reaction

    T cell recovery after irradiation-induced lymphopenia. B6 mice, Fas fl/fl lck-cre, and Fas fl/fl mice were irradiated with 6 Gy, CD3 T cells in the spleen were counted and analyzed by flow cytometry. Approximately 3–4 mice per group per time point (5-time points total) were analyzed.

    Journal: Frontiers in Immunology

    Article Title: Allergen Exposure in Lymphopenic Fas-Deficient Mice Results in Persistent Eosinophilia Due to Defects in Resolution of Inflammation

    doi: 10.3389/fimmu.2018.02395

    Figure Lengend Snippet: T cell recovery after irradiation-induced lymphopenia. B6 mice, Fas fl/fl lck-cre, and Fas fl/fl mice were irradiated with 6 Gy, CD3 T cells in the spleen were counted and analyzed by flow cytometry. Approximately 3–4 mice per group per time point (5-time points total) were analyzed.

    Article Snippet: Antibodies and flow cytometry Anti-mouse CD3 (clone 17A2; BD Biosciences, San Diego, CA), anti-mouse CCR3 (clone 83101.111; R & D Systems, Minneapolis, MN), and anti-mouse Ly6G (GR1, BD Biosciences) antibody were used for flow of bronchial alveolar lavage (BAL) T cells (CD3+ side scatter low) and eosinophils (CCR3+ Ly6G− and side scatter high).

    Techniques: Irradiation, Mouse Assay, Flow Cytometry, Cytometry

    Ectopic Sestrin2 expression in M1 macrophages attenuates post-MI inflammation and favors tissue repair in hearts. (A) Macrophage abundance in infarcted myocardium at day 3 after myocardial infarction (MI) (day 4 after Clophosome ® treatment). Ctrl: un-pretreated mice. Depletion: Clophosome ® injection. Depletion + M (L-C): Clophosome ® injection followed by transfer of M1 macrophages transduced with control lentivirus. Depletion + M (L-S): Clophosome ® injection followed by transfer of M1 macrophages transduced with SESN2 lentiviral activation particles. Numbers in the plots are proportions of gated cells populations. This is a representative of two independent experiments. (B,C) mRNA abundance of indicated cytokines and chemokines in infarcted myocardium. (D,E) Percentage of Gr-1 + neutrophils in the whole cardiac cells at day 3 after MI. Representative flow cytometry dot plots are shown in (D) , and statistics is shown in (E) . (F) Masson staining at day 7 after MI. Left panel: representative images. Right panel: statistics of collagen volume fraction (CVF). N = 4–5 per group. * p

    Journal: Frontiers in Immunology

    Article Title: Sestrin2 Suppresses Classically Activated Macrophages-Mediated Inflammatory Response in Myocardial Infarction through Inhibition of mTORC1 Signaling

    doi: 10.3389/fimmu.2017.00728

    Figure Lengend Snippet: Ectopic Sestrin2 expression in M1 macrophages attenuates post-MI inflammation and favors tissue repair in hearts. (A) Macrophage abundance in infarcted myocardium at day 3 after myocardial infarction (MI) (day 4 after Clophosome ® treatment). Ctrl: un-pretreated mice. Depletion: Clophosome ® injection. Depletion + M (L-C): Clophosome ® injection followed by transfer of M1 macrophages transduced with control lentivirus. Depletion + M (L-S): Clophosome ® injection followed by transfer of M1 macrophages transduced with SESN2 lentiviral activation particles. Numbers in the plots are proportions of gated cells populations. This is a representative of two independent experiments. (B,C) mRNA abundance of indicated cytokines and chemokines in infarcted myocardium. (D,E) Percentage of Gr-1 + neutrophils in the whole cardiac cells at day 3 after MI. Representative flow cytometry dot plots are shown in (D) , and statistics is shown in (E) . (F) Masson staining at day 7 after MI. Left panel: representative images. Right panel: statistics of collagen volume fraction (CVF). N = 4–5 per group. * p

    Article Snippet: Flow Cytometry Analysis Anti-mouse antibodies were used: APC anti-CD3 (17A2), APC anti-B220 (RA3-6B2), APC anti-Gr1 (RB6-8C5), APC anti-CD90 (OX-7), FITC anti-CD11b (M1/70), PE anti-F4/80 (T45-2342), and PE-Cy7 anti-Ly-6C (AL-21) were purchased from BD Biosciences.

    Techniques: Expressing, Mouse Assay, Injection, Transduction, Activation Assay, Flow Cytometry, Cytometry, Staining

    Ectopic Sestrin2 expression in M1 macrophages inhibits pro-inflammatory response in vitro . (A) Expression of CD80 and CD163 on in vitro cultured M1 and M2 monocyte-derived macrophages. Numbers in the quadrants are proportions of corresponding subpopulations. This is a representative of two independent experiments. (B) Lentiviral transduction efficiency of cultured macrophages is detected with flow cytometry. Note that here macrophages were not transduced with GFP-free SESN2 lentiviral activation particles. Instead, they were transduced by copGFP lentiviral particles containing GFP sequence. Numbers in the histogram are percentages of GFP + cells. (C) Sestrin2 protein levels in M1 and M2 macrophages after lentiviral transduction. L-C: control lentivirus not inducing Sestrin2 expression. L-S: SESN2 lentiviral activation particles. This is a representative of three independent experiments. (D–F) mRNA abundance of TNF-α, IL-1β, and IL-10 in lentivirus-transduced M1 and M2 macrophages with or without lipopolysaccharide (LPS) stimulation. M1 L-C: M1 macrophages transduced with control lentivirus. M1 L-S: M1 macrophages transduced with SESN2 lentiviral activation particles. M2 L-C: M2 macrophages transduced with control lentivirus. M2 L-S: M2 macrophages transduced with SESN2 lentiviral activation particles. N = 6 per group. * p

    Journal: Frontiers in Immunology

    Article Title: Sestrin2 Suppresses Classically Activated Macrophages-Mediated Inflammatory Response in Myocardial Infarction through Inhibition of mTORC1 Signaling

    doi: 10.3389/fimmu.2017.00728

    Figure Lengend Snippet: Ectopic Sestrin2 expression in M1 macrophages inhibits pro-inflammatory response in vitro . (A) Expression of CD80 and CD163 on in vitro cultured M1 and M2 monocyte-derived macrophages. Numbers in the quadrants are proportions of corresponding subpopulations. This is a representative of two independent experiments. (B) Lentiviral transduction efficiency of cultured macrophages is detected with flow cytometry. Note that here macrophages were not transduced with GFP-free SESN2 lentiviral activation particles. Instead, they were transduced by copGFP lentiviral particles containing GFP sequence. Numbers in the histogram are percentages of GFP + cells. (C) Sestrin2 protein levels in M1 and M2 macrophages after lentiviral transduction. L-C: control lentivirus not inducing Sestrin2 expression. L-S: SESN2 lentiviral activation particles. This is a representative of three independent experiments. (D–F) mRNA abundance of TNF-α, IL-1β, and IL-10 in lentivirus-transduced M1 and M2 macrophages with or without lipopolysaccharide (LPS) stimulation. M1 L-C: M1 macrophages transduced with control lentivirus. M1 L-S: M1 macrophages transduced with SESN2 lentiviral activation particles. M2 L-C: M2 macrophages transduced with control lentivirus. M2 L-S: M2 macrophages transduced with SESN2 lentiviral activation particles. N = 6 per group. * p

    Article Snippet: Flow Cytometry Analysis Anti-mouse antibodies were used: APC anti-CD3 (17A2), APC anti-B220 (RA3-6B2), APC anti-Gr1 (RB6-8C5), APC anti-CD90 (OX-7), FITC anti-CD11b (M1/70), PE anti-F4/80 (T45-2342), and PE-Cy7 anti-Ly-6C (AL-21) were purchased from BD Biosciences.

    Techniques: Expressing, In Vitro, Cell Culture, Derivative Assay, Transduction, Flow Cytometry, Cytometry, Activation Assay, Sequencing

    Sestrin2 inhibits mTORC1 signal pathway in M1 macrophages. (A) mTOR phosphorylation in transferred M1 macrophages. M1 macrophages were transduced with lentivirus and transferred into recipient mice as in Figure 5 . These macrophages were then sorted using flow cytometry from infarcted myocardium at day 3 after myocardial infarction (MI) for detection of mTORC1 activation status. L-C: M1 macrophages transduced with control lentivirus. L-S: M1 macrophages transduced with SESN2 lentiviral activation particles. #1 to #3 indicate three mice. (B) mTOR phosphorylation in in vitro cultured M1 macrophages after lentiviral transduction. (C) 4EBP1 phosphorylation in transferred M1 macrophages at day 3 after MI. N = 3 per group. (D) mTOR phosphorylation in in vitro cultured M1 macrophages in the presence or absence of 1-h MHY1485 treatment. L-C: M1 macrophages transduced with control lentivirus. L-S: M1 macrophages transduced with SESN2 lentiviral activation particles. This is a representative image of two independent experiments. (E–G) mRNA abundance of indicated cytokines in in vitro cultured M1 macrophages. LPS: LPS stimulation. M + LPS: MHY1485 pretreatment followed by LPS stimulation. N = 5 per group. * p

    Journal: Frontiers in Immunology

    Article Title: Sestrin2 Suppresses Classically Activated Macrophages-Mediated Inflammatory Response in Myocardial Infarction through Inhibition of mTORC1 Signaling

    doi: 10.3389/fimmu.2017.00728

    Figure Lengend Snippet: Sestrin2 inhibits mTORC1 signal pathway in M1 macrophages. (A) mTOR phosphorylation in transferred M1 macrophages. M1 macrophages were transduced with lentivirus and transferred into recipient mice as in Figure 5 . These macrophages were then sorted using flow cytometry from infarcted myocardium at day 3 after myocardial infarction (MI) for detection of mTORC1 activation status. L-C: M1 macrophages transduced with control lentivirus. L-S: M1 macrophages transduced with SESN2 lentiviral activation particles. #1 to #3 indicate three mice. (B) mTOR phosphorylation in in vitro cultured M1 macrophages after lentiviral transduction. (C) 4EBP1 phosphorylation in transferred M1 macrophages at day 3 after MI. N = 3 per group. (D) mTOR phosphorylation in in vitro cultured M1 macrophages in the presence or absence of 1-h MHY1485 treatment. L-C: M1 macrophages transduced with control lentivirus. L-S: M1 macrophages transduced with SESN2 lentiviral activation particles. This is a representative image of two independent experiments. (E–G) mRNA abundance of indicated cytokines in in vitro cultured M1 macrophages. LPS: LPS stimulation. M + LPS: MHY1485 pretreatment followed by LPS stimulation. N = 5 per group. * p

    Article Snippet: Flow Cytometry Analysis Anti-mouse antibodies were used: APC anti-CD3 (17A2), APC anti-B220 (RA3-6B2), APC anti-Gr1 (RB6-8C5), APC anti-CD90 (OX-7), FITC anti-CD11b (M1/70), PE anti-F4/80 (T45-2342), and PE-Cy7 anti-Ly-6C (AL-21) were purchased from BD Biosciences.

    Techniques: Transduction, Mouse Assay, Flow Cytometry, Cytometry, Activation Assay, In Vitro, Cell Culture

    Sestrin2 is expressed in cardiac macrophages after myocardial infarction (MI). (A) Representative flow cytometry dot plots showing cardiac macrophage subpopulations in infarcted myocardium at day 1, day 3, day 5, and day 7 after MI. Numbers in the quadrants are proportions of corresponding subpopulations. Sham: Sham-operated animal. This is a representative of three independent experiments. (B) Sestrin2 protein levels in total cardiac macrophages (upper panel) and blood monocytes (lower panel). (C) Statistics for (B) . N = 3 per group. * p

    Journal: Frontiers in Immunology

    Article Title: Sestrin2 Suppresses Classically Activated Macrophages-Mediated Inflammatory Response in Myocardial Infarction through Inhibition of mTORC1 Signaling

    doi: 10.3389/fimmu.2017.00728

    Figure Lengend Snippet: Sestrin2 is expressed in cardiac macrophages after myocardial infarction (MI). (A) Representative flow cytometry dot plots showing cardiac macrophage subpopulations in infarcted myocardium at day 1, day 3, day 5, and day 7 after MI. Numbers in the quadrants are proportions of corresponding subpopulations. Sham: Sham-operated animal. This is a representative of three independent experiments. (B) Sestrin2 protein levels in total cardiac macrophages (upper panel) and blood monocytes (lower panel). (C) Statistics for (B) . N = 3 per group. * p

    Article Snippet: Flow Cytometry Analysis Anti-mouse antibodies were used: APC anti-CD3 (17A2), APC anti-B220 (RA3-6B2), APC anti-Gr1 (RB6-8C5), APC anti-CD90 (OX-7), FITC anti-CD11b (M1/70), PE anti-F4/80 (T45-2342), and PE-Cy7 anti-Ly-6C (AL-21) were purchased from BD Biosciences.

    Techniques: Flow Cytometry, Cytometry

    LINC00982 and LL22NC03-N14H11.1 have opposite roles in the proliferation of gastric cancer cells. (A) Reverse transcription-quantitative polymerase chain reaction revealed that the expression of LINC00982 in the LINC00982-overexpressed SGC-7901 cell line (left) and the expression of LL22NC03-N14H11.1 in the LL22NC03-N14H11.1-down-regulated SGC-7901 cell line (right). (B) The MTT assay revealed the proliferative status of the SGC-7901 cell line after LINC00982 being overexpressed or LL22NC03-N14H11.1 being downregulated. (C) Flow cytometric analysis of the expression of Ki-67 in the SGC-7901 cell line after LINC00982 was overexpressed or LL22NC03-N14H11.1 was downregulated. All the experiments were replicated a minimum of 3. The error bars represent the standard deviation. *P

    Journal: Oncology Letters

    Article Title: Comprehensive bioinformatics analysis of lncRNAs in gastric cancer

    doi: 10.3892/ol.2018.9707

    Figure Lengend Snippet: LINC00982 and LL22NC03-N14H11.1 have opposite roles in the proliferation of gastric cancer cells. (A) Reverse transcription-quantitative polymerase chain reaction revealed that the expression of LINC00982 in the LINC00982-overexpressed SGC-7901 cell line (left) and the expression of LL22NC03-N14H11.1 in the LL22NC03-N14H11.1-down-regulated SGC-7901 cell line (right). (B) The MTT assay revealed the proliferative status of the SGC-7901 cell line after LINC00982 being overexpressed or LL22NC03-N14H11.1 being downregulated. (C) Flow cytometric analysis of the expression of Ki-67 in the SGC-7901 cell line after LINC00982 was overexpressed or LL22NC03-N14H11.1 was downregulated. All the experiments were replicated a minimum of 3. The error bars represent the standard deviation. *P

    Article Snippet: Anti-mouse-ki-67-PE flow cytometric antibody (eBioscience; Thermo Fisher Scientific, Inc.) was purchased from eBioscience.

    Techniques: Real-time Polymerase Chain Reaction, Expressing, MTT Assay, Flow Cytometry, Standard Deviation

    ZIKV infection of unstimulated and decidualized T-HESC cell line. (A) T-HESC (left) and decidualized (d)T-HESC (right) were stained with anti-dsRNA or Flavi E mAbs whereas the nuclei were stained with Hoechst. Scale bar: 20 µm. (B) Surface expression of AXL (red) and MER (blue) in T-HESC (left) and dT-HESC (right) was determined by flow cytometry. The histograms of one experiment representative of 3 independently performed are shown. Double immunostaining for Flavi E and calreticulin (C) or vimentin (D) in T-HESC (left) and dT-HESC (right) either uninfected or infected with MR766 at 72 h post-infection; Hoechst was used to stain nuclei. Scale bar: 10 µm.

    Journal: bioRxiv

    Article Title: Human Endometrial Stromal Cells Are Highly Permissive To Productive Infection by Zika Virus

    doi: 10.1101/077305

    Figure Lengend Snippet: ZIKV infection of unstimulated and decidualized T-HESC cell line. (A) T-HESC (left) and decidualized (d)T-HESC (right) were stained with anti-dsRNA or Flavi E mAbs whereas the nuclei were stained with Hoechst. Scale bar: 20 µm. (B) Surface expression of AXL (red) and MER (blue) in T-HESC (left) and dT-HESC (right) was determined by flow cytometry. The histograms of one experiment representative of 3 independently performed are shown. Double immunostaining for Flavi E and calreticulin (C) or vimentin (D) in T-HESC (left) and dT-HESC (right) either uninfected or infected with MR766 at 72 h post-infection; Hoechst was used to stain nuclei. Scale bar: 10 µm.

    Article Snippet: Flow Cytometry Mouse anti-human AXL mAb (clone # 108724, MAB154) and mouse anti-human MER mAb (clone # 125518, MAB8912) was purchased from R & D Systems.

    Techniques: Infection, Staining, Expressing, Flow Cytometry, Double Immunostaining

    CD151 deficiency does not alter basal phenotype of cultured BMMCs. ( A ) CD151 deficiency does not affect growth dynamics of mast cell culture. WT and CD151 −/− BMMCs were cultured in IL-3–conditioned media for 8 wk and the total numbers of live cells in culture were counted at each time point indicated ( left ). Flow cytometry for CD151 expression on peritoneal mast cells (flow cytometry chart) ( middle ), qPCR detection of CD151 mRNA in WT and CD151 −/− BMMCs (bar graph, right ), and Western blot analysis of total-protein lysates for CD151 expression (Western blot, right ) all confirm constitutive CD151 expression in WT mast cells. WT peritoneal mast cells are shown as gray-filled histogram with dotted line and negative control as transparent histogram with dotted line. In immunoblotting, actin was used as a loading control. ( B ) Five-week-old BMMCs from WT and CD151 −/− mice were stained with toluidine blue and images were obtained with an original magnification of ×100. Flow cytometry analysis of FcεRI and c-Kit surface expression and purity of WT and CD151 −/− BMMC cultures. All data are representative of three independent experiments. Data are represented as mean ± SEM. * p

    Journal: Journal of immunology (Baltimore, Md. : 1950)

    Article Title: Tetraspanin CD151 Is a Negative Regulator of FcεRI-Mediated Mast Cell Activation

    doi: 10.4049/jimmunol.1302874

    Figure Lengend Snippet: CD151 deficiency does not alter basal phenotype of cultured BMMCs. ( A ) CD151 deficiency does not affect growth dynamics of mast cell culture. WT and CD151 −/− BMMCs were cultured in IL-3–conditioned media for 8 wk and the total numbers of live cells in culture were counted at each time point indicated ( left ). Flow cytometry for CD151 expression on peritoneal mast cells (flow cytometry chart) ( middle ), qPCR detection of CD151 mRNA in WT and CD151 −/− BMMCs (bar graph, right ), and Western blot analysis of total-protein lysates for CD151 expression (Western blot, right ) all confirm constitutive CD151 expression in WT mast cells. WT peritoneal mast cells are shown as gray-filled histogram with dotted line and negative control as transparent histogram with dotted line. In immunoblotting, actin was used as a loading control. ( B ) Five-week-old BMMCs from WT and CD151 −/− mice were stained with toluidine blue and images were obtained with an original magnification of ×100. Flow cytometry analysis of FcεRI and c-Kit surface expression and purity of WT and CD151 −/− BMMC cultures. All data are representative of three independent experiments. Data are represented as mean ± SEM. * p

    Article Snippet: Allophycocyanin-conjugated anti-mouse CD151 flow cytometry Ab was from R & D Systems (Minneapolis, MN).

    Techniques: Cell Culture, Flow Cytometry, Cytometry, Expressing, Real-time Polymerase Chain Reaction, Western Blot, Negative Control, Mouse Assay, Staining

    Extended validation of candidate genes displaying stronger dropout effects in STAG2 -null versus wild-type cells. (A) Cas9-GFP–expressing KBM-7 clonal cell line (B4) was lentivirally transduced with a validated sgRNA targeting STAG2 , and several subclones were isolated and characterized for STAG2 knockout by immunoblotting. (B) For each candidate gene, multiple sgRNAs scoring in the primary screen (2 or 3, as indicated) were tested in parallel competitive proliferation assays in STAG2 –wild-type (B4, blue lines) and STAG2 -mutant (c9, red lines) cell lines used in the screen and in an independent STAG2 -mutant clone (c11, orange lines). Cells were transduced with lentiviral vectors co-expressing mCherry and an sgRNA targeting the indicated genes, and the fraction of mCherry+ cells was monitored over time using flow cytometry. Plotted are averages of 2–3 independent sgRNAs analyzed for each gene; error bars represent standard deviations. Note that knockout of most candidates (e.g., BUB1, PAXIP1, PDCD5, PP2R1A, and SKA3) was deleterious in both STAG2–wild-type and STAG2-mutant cells, with some differences in depletion dynamics. In other instances, knockout effects turned out to be specific without being strongly detrimental in STAG2-mutant cells (e.g., CHMP7, FLI1, and SKI), and in some cases, they could not be confirmed in an independent STAG2-null clone (c11, e.g., DNAJC9 and FANCI), indicating that clonal artifacts remain an issue even when starting out from a single-cell–derived clone.

    Journal: Life Science Alliance

    Article Title: STAG1 vulnerabilities for exploiting cohesin synthetic lethality in STAG2-deficient cancers

    doi: 10.26508/lsa.202000725

    Figure Lengend Snippet: Extended validation of candidate genes displaying stronger dropout effects in STAG2 -null versus wild-type cells. (A) Cas9-GFP–expressing KBM-7 clonal cell line (B4) was lentivirally transduced with a validated sgRNA targeting STAG2 , and several subclones were isolated and characterized for STAG2 knockout by immunoblotting. (B) For each candidate gene, multiple sgRNAs scoring in the primary screen (2 or 3, as indicated) were tested in parallel competitive proliferation assays in STAG2 –wild-type (B4, blue lines) and STAG2 -mutant (c9, red lines) cell lines used in the screen and in an independent STAG2 -mutant clone (c11, orange lines). Cells were transduced with lentiviral vectors co-expressing mCherry and an sgRNA targeting the indicated genes, and the fraction of mCherry+ cells was monitored over time using flow cytometry. Plotted are averages of 2–3 independent sgRNAs analyzed for each gene; error bars represent standard deviations. Note that knockout of most candidates (e.g., BUB1, PAXIP1, PDCD5, PP2R1A, and SKA3) was deleterious in both STAG2–wild-type and STAG2-mutant cells, with some differences in depletion dynamics. In other instances, knockout effects turned out to be specific without being strongly detrimental in STAG2-mutant cells (e.g., CHMP7, FLI1, and SKI), and in some cases, they could not be confirmed in an independent STAG2-null clone (c11, e.g., DNAJC9 and FANCI), indicating that clonal artifacts remain an issue even when starting out from a single-cell–derived clone.

    Article Snippet: 4 d postinfection, the transduction efficacy was determined using antibody staining for Thy1.1 (APC anti-mouse CD90.1, 202526; BioLegend) and flow cytometry analysis.

    Techniques: Expressing, Transduction, Isolation, Knock-Out, Mutagenesis, Flow Cytometry, Derivative Assay

    Genome-wide CRISPR screens in isogenic KBM-7 cell lines identify STAG1 as the most selective synthetic-lethal dependency in STAG2 -mutant cells. (A) Schematic of cell line engineering and genome-wide CRISPR screening. Haploid KBM-7 cells were sequentially transduced with indicated lentiviral vectors, Cas9-GFP–expressing cells were single-cell sorted, and derived clones characterized for homogenous and effective CRISPR editing. The selected clone was lentivirally transduced with a validated sgRNA-targeting STAG2 , and several subclones were isolated and characterized for STAG2 knockout. One STAG2 knockout clone was selected (“c9”) and screened side-by-side with the parental STAG2 –wild-type clone (“B4”) using a second-generation genome-wide sgRNA library. (B) Gene-level dropout effects in wild-type and STAG2 -null KBM-7 cells. Shown are log 2 fold changes between the end point of triplicate screens and the sgRNA library (analyzed using MAGeCK v0.5.8). A set of previously described generally essential genes ( Wang et al, 2019 ) are highlighted in red. (C) Analysis of differential effects in STAG2 -null versus STAG2 –wil-type cells (MAGeCK v0.5.8), revealing STAG1 as the most prominent and only significant synthetic lethal interaction. (D) Competitive proliferations assays in STAG2 –wild-type (B4, blue lines) and STAG2 -mutant (c9, red lines) cell lines used in the screen and in an independent STAG2 -mutant clone (c11, orange lines). Cells were transduced with a lentiviral vector co-expressing mCherry and the indicated sgRNAs, and the fraction of mCherry+ cells was monitored over time using flow cytometry.

    Journal: Life Science Alliance

    Article Title: STAG1 vulnerabilities for exploiting cohesin synthetic lethality in STAG2-deficient cancers

    doi: 10.26508/lsa.202000725

    Figure Lengend Snippet: Genome-wide CRISPR screens in isogenic KBM-7 cell lines identify STAG1 as the most selective synthetic-lethal dependency in STAG2 -mutant cells. (A) Schematic of cell line engineering and genome-wide CRISPR screening. Haploid KBM-7 cells were sequentially transduced with indicated lentiviral vectors, Cas9-GFP–expressing cells were single-cell sorted, and derived clones characterized for homogenous and effective CRISPR editing. The selected clone was lentivirally transduced with a validated sgRNA-targeting STAG2 , and several subclones were isolated and characterized for STAG2 knockout. One STAG2 knockout clone was selected (“c9”) and screened side-by-side with the parental STAG2 –wild-type clone (“B4”) using a second-generation genome-wide sgRNA library. (B) Gene-level dropout effects in wild-type and STAG2 -null KBM-7 cells. Shown are log 2 fold changes between the end point of triplicate screens and the sgRNA library (analyzed using MAGeCK v0.5.8). A set of previously described generally essential genes ( Wang et al, 2019 ) are highlighted in red. (C) Analysis of differential effects in STAG2 -null versus STAG2 –wil-type cells (MAGeCK v0.5.8), revealing STAG1 as the most prominent and only significant synthetic lethal interaction. (D) Competitive proliferations assays in STAG2 –wild-type (B4, blue lines) and STAG2 -mutant (c9, red lines) cell lines used in the screen and in an independent STAG2 -mutant clone (c11, orange lines). Cells were transduced with a lentiviral vector co-expressing mCherry and the indicated sgRNAs, and the fraction of mCherry+ cells was monitored over time using flow cytometry.

    Article Snippet: 4 d postinfection, the transduction efficacy was determined using antibody staining for Thy1.1 (APC anti-mouse CD90.1, 202526; BioLegend) and flow cytometry analysis.

    Techniques: Genome Wide, CRISPR, Mutagenesis, Transduction, Expressing, Derivative Assay, Clone Assay, Isolation, Knock-Out, Plasmid Preparation, Flow Cytometry