fluorescein isothiocyanate Search Results


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  • 99
    Thermo Fisher fitc
    Interaction of Candid 1 with VGCC subunits on the cell surface. (a) Cells treated with the indicated siRNAs and incubated with <t>FITC-labeled</t> Candid 1. Shown is a representative <t>FACs</t> plot and below the graph, the average MFI of the peaks averaged from 2
    Fitc, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 11713 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore fitc
    Activation of the m-calpain isoform in the ACT-treated cells. Western blot of the cytosolic fraction isolated from J774A.1 cells treated with ACT (35 nM) or from cells transfected with anti m-calpain siRNA and treated with ACT (35 nM) (A). Kinetics of the proteolytic cleavage of the FRET-based calpain specific substrate for ACT-treated cells (35 nM toxin), and cells in which m-calpain expression has been silenced by transfection with the corresponding siRNA, or mock cells transfected with scrambled control siRNA, or cells pre-incubated with SAJ6017 (78 nM), a specific m-calpain inhibitor. The figure is a representative experiment from three similar assays performed under identical conditions (B). Assay of calpain activity using <t>FITC-labelled</t> <t>α-casein</t> as substrate. FITC-α-casein hydrolysis by calpain is determined from in J774A.1 cell extracts incubated with FITC-labelled α-casein in the presence or absence of 1 mM CaCl 2 . Protease activity was quantified in 10% polyacrylamide gel by determining fluorescence intensity of the non-hydrolysed FITC-α-casein band. Non-digested FITC-α-casein and cell extracts without calcium were used as negative controls (C).
    Fitc, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 8829 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore fluorescein isothiocyanate
    Immunohistochemistry for T. gondii detection using polyclonal IgY anti-STAg antibodies. Paraffin-embedded brain sections of mice chronically infected with ME-49 strain were incubated with IgY anti-STAg and rabbit IgG anti-IgY conjugated to fluorescein <t>isothiocyanate</t> (FITC). ( A and B ), segmented parasitophorus vacuoles were detected into host cell cytoplasm around the DAPI-stained nucleus (blue). ( C and D ), IgY anti-STAg antibodies strongly recognized antigens on outer walls of tissue cysts and free tachyzoites ( C , arrow).
    Fluorescein Isothiocyanate, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 4454 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Becton Dickinson fluorescein isothiocyanate
    Microbubble (MB) binding to receptor expression. Correlation of MB EGFR (A), MB CD147 (B), and MB DUAL (C) binding to receptor expression was assessed in 4 head and neck cancer cell lines (SCC-1, SCC-5, FaDu, and Cal27), 2 head and neck cancer CD147 knock-down cell lines (SCC-1/SiE-711 and SCC-5/SiE-711), and normal dermal fibroblasts (NDFs). FITC indicates fluorescein <t>isothiocyanate.</t> MB IgG , MB EGFR , MB CD147 , and MB DUAL indicate IgG-, anti-EGFR–, and anti-CD147–targeted MBs and dual-conjugated MBs, respectively.
    Fluorescein Isothiocyanate, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 93/100, based on 3902 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Becton Dickinson fluorescein isothiocyanate fitc
    Percentage of Th1, Th2, and Th0 cells in nonpregnant women, normal pregnant women, and preeclamptic patients. Peripheral blood mononuclear cells were stained with Peridinin chlorophyll protein (PerCP)-conjugated CD4 monoclonal antibody, <t>fluorescein-isothiocyanate</t> <t>(FITC)-conjugated</t> anti IFN-γ monoclonal antibody and PE-conjugated anti-interleukin (IL)-4 monoclonal antibody, as described in Materials and Methods. A gate was set on the CD4 + lymphocytes by characteristic forward and side scatter parameters (R1 and R2). The FITC (anti-interferon (IFN)-γ) and PE (anti-IL-4) fluorescence data were analysed. Isotype muched fluorochrome-conjugated IgG1 and IgG2a were used as control. The horizontal bar shows FITC fluorescence and the vertical bar shows PE fluorescence. Fluorescence intensity is determined on a logarithmic scale.
    Fluorescein Isothiocyanate Fitc, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 92/100, based on 4063 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore fluorescein isothiocyanate dextran
    UDP inhibits C5a-stimulated macropinocytosis. BMDM were stimulated with or without C5a (0, 0.3, 0.75, and 2.5 n m ) in the presence or absence of UDP (0, 0.5, and 2.5 μ m ) in assays of macropinocytosis. Uptake of extracellular fluorescein <t>isothiocyanate-dextran</t>
    Fluorescein Isothiocyanate Dextran, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 1138 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Thermo Fisher fluorescein isothiocyanate
    DSPS alters cell cycle progression and promotes apoptosis. ( a ) MDA‐MB‐231 cells treated with various concentrations of DSPS were stained with PI, and the cells were analyzed by flow cytometry to determine cell cycle progression. ( b ) Quantification of the percentage of cells in G1, S, and G2/M phases of cells treated as in panel A. 0 mg/mL, 0.5 mg/mL, 5 mg/mL. ( c–f ) MDA‐MB‐231 cells treated with various concentrations of DSPS and stained with Annexin V fluorescein <t>isothiocyanate</t> (FITC)/propidium iodide ( c ); the percentage of total apoptotic cells ( d ), cells in early apoptosis ( e ), and cells in late apoptosis ( f ) were quantified.
    Fluorescein Isothiocyanate, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 3267 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Becton Dickinson annexin v fluorescein isothiocyanate
    3-MA significantly increased the apoptotic rate under hypoxic conditions. Apoptosis was detected by a Annexin V-fluorescein <t>isothiocyanate</t> assay. (A) Flow cytometry analysis of apoptosis. Q2 represented the early apoptotic cells. Q4 represented the late apoptotic cells. Apoptotic cells from each group were detected by <t>Annexin</t> V and propidium iodide. (B) The proportion of apoptotic cells from each group was quantified. Data are presented as the mean ± standard deviation. *P
    Annexin V Fluorescein Isothiocyanate, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 92/100, based on 3114 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Becton Dickinson annexin v fluorescein isothiocyanate fitc
    Effect of different concentrations of udenafil for 24 h on apoptosis in VEC and VSMC. Distributions of cells treated with different concentrations of udenafil displayed as dot plots: Viable cells (fluorescein <t>isothiocyanate</t> (FITC˜)/propidium iodide (PĨ)), apoptotic cells <t>(FITC+/PĨ),</t> secondary necrotic cells (FITC+/PI+). (a and f) Cells incubated with 0 μmol/l udenafil. (b and g) Cells incubated with 1 μmol/l udenafil. (c and h) Cells incubated with 10 μmol/l udenafil. (d and i) Cells incubated with 100 μmol/l udenafil. (e and j) Cells incubated with 1 mmol/l udenafil. A minimum of 10,000 events was counted per sample
    Annexin V Fluorescein Isothiocyanate Fitc, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 94/100, based on 1872 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Agilent technologies fluorescein isothiocyanate
    Immunofluorescence analysis of virus maturation. HFF were infected with RV wt AD169 or RV del UL35 at an MOI of 0.2. Cells were fixed at indicated time points (24, 72, and 120 hpi) and stained with monoclonal antibodies directed against ppUL82 (CMV355), ppUL83 (28-77), ppUL99 (41-18), and gpUL55 (27-287), as indicated at the top. Goat anti-mouse secondary antibody labeled with fluorescein <t>isothiocyanate</t> was used for detection. The nuclei were stained with DAPI. See the text for details.
    Fluorescein Isothiocyanate, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 92/100, based on 593 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Becton Dickinson annexin v fluorescein isothiocyanate fitc apoptosis detection kit
    Cholangiocarcinoma QBC939 cells <t>apoptosis</t> induced by solamargine. (A) Apoptosis of QBC939 cells induced by treatment with different concentrations of solamargine for 24 h, stained using Annexin <t>V-FITC/PI</t> and determined using flow cytometry. (B) Statistical data of QBC939 cell apoptosis. Apoptosis rate contains the early apoptosis rate and late apoptosis rate. *P
    Annexin V Fluorescein Isothiocyanate Fitc Apoptosis Detection Kit, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 94/100, based on 1115 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Becton Dickinson fluorescein isothiocyanate fitc conjugated annexin v
    Interferon (IFN)-γ in combination with TNF-α induces cell death in mouse lung carcinoma cells. L929 fibroblasts and Lewis lung carcinoma (LLC) cancer cells were cultured for 24 h before starting treatment with recombinant cytokines. Both L929 and LLC cells were (A,B) left untreated for 48 h and used as negative controls or (C) L929 cells were treated with 50 ng/mL TNF-α for 48 h and used as a positive control. LLC cell viability was retained after treatment with (D) 50 ng/mL TNF-α or (E) 100 ng/mL IFN-γ and after (F) simultaneous treatment with both 50 ng/mL TNF-α and 100 ng/mL IFN-γ for 24 h. Cell death was induced in LLC cells after (G) simultaneous treatment with 100 ng/mL of IFN-γ in combination with 50 ng/mL of TNF-α for 48 h and after (H) LLC pretreatment with 100 ng/mL of IFN-γ for 24 h followed by treatment with TNF-α for 24 h. The resulting cell monolayers were analyzed using bright-field microscopy (scale bar, 50 µM). Cell death was determined using flow cytometry analysis after cell staining with annexin <t>V-FITC</t> (depicted on the x -axis) and propidium iodide (PI, depicted on the y -axis). Annexin V-positive/PI-negative cells were regarded as apoptotic, whereas annexin V-positive/PI-positive cells were regarded as necrotic. Experiment was performed in duplicates and repeated two times with standard error not exceeding 10% between independent experiments. One representative experiment is shown, where the percentages of the four distinct cell populations are averages of duplicates with SEM
    Fluorescein Isothiocyanate Fitc Conjugated Annexin V, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 92/100, based on 672 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Santa Cruz Biotechnology fluorescein isothiocyanate
    M1 macrophages activate HES1 expression and markers of differentiation in epithelial cells. HT29 cells (a) or Caco-2 cells (b) at pre-confluence were co-cultured (24h) with M1 or M2 macrophages (stained with CD18 and fluorescein <t>isothiocyanate).</t> In some
    Fluorescein Isothiocyanate, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 94/100, based on 590 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore fitc labeled phalloidin
    Effects of FRAX486 and IPA3 on actin organization in WPMY-1 cells. Actin filaments were visualized by staining with <t>FITC-coupled</t> <t>phalloidin,</t> after incubation of WPMY-1 cells with DMSO, FRAX486 (1–10 μM), or IPA3 (1–10 μM) for 24 h. Shown are representative images from series with 5 independent experiments, with similar results.
    Fitc Labeled Phalloidin, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 761 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    fitc  (Abcam)
    99
    Abcam fitc
    Confirmation of putative receptor-binding sites on the ligands using synthetic analogues by immunocytochemistry. Interaction of synthetic analogues of putative receptor-binding sites with cultured <t>hBMECs.</t> The interaction was detected with <t>streptavidin-FITC</t> conjugate. Nuclei are stained with DAPI. rDIII and rNadA (positive control) – whole recombinant ligands were incubated with hBMECs. DIII-1 – GTTYGVCSK-biotin; DIII-2 – VLIELEPPFGDSYIVVGRK-biotin; NadA-1 – AATVAIVAAYNNGQEINGFKAGETIYDIGEDGTITQK-biotin; NadA-2 – LADTDAALADTDAALDETTNALNKLGENITTFAEETK-biotin. Negative control – synthetic peptides were excluded from the assay.
    Fitc, supplied by Abcam, used in various techniques. Bioz Stars score: 99/100, based on 1845 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Pharmingen fluorescein isothiocyanate
    Expression of surface molecules on NRS1 and 4-1BBL-transfected NRS1 cells. Parental NRS1 and 4-1BBL-transfected NRS1 (clone 5.13) cells were stained with either fluorescein <t>isothiocyanate-</t> or phycoerythrin-conjugated anti-4-1BBL, anti-CD80, anti-CD86, anti-CD54, anti-H-2K k and anti-I-A k , or an appropriate fluorochrome-conjugated immunoglobulin control. Samples were analysed by flow cytometry. Data are displayed as histograms (4-decade scale) with the immunoglobulin control as the lighter line.
    Fluorescein Isothiocyanate, supplied by Pharmingen, used in various techniques. Bioz Stars score: 94/100, based on 524 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Agilent technologies fluorescein isothiocyanate fitc
    Intracellular distribution of U14. (A) Time course IFA of HST-infected Molt-3 cells harvested at the indicated time points (hpi). Acetone-fixed cells were incubated with anti-U14 and anti-p53 (DO-1) MAbs and then labeled with <t>FITC-conjugated</t> goat anti-mouse immunoglobulin G. (B) Confocal microscopy of HST-infected Molt-3 cells after 18 h. Anti-p53 goat PAb (FL393) and anti-U14 MAb were reacted with FITC- and tetramethyl rhodamine <t>isothiocyanate-labeled</t> secondary antibodies, respectively. Arrows and arrowheads indicate the cytoplasmic and nuclear dots, respectively. (C) 293 cells were transfected with the U14 expression plasmid, pEF-U14, and subjected to IFA at 24 h posttransfection. U14 was labeled with FITC-conjugated secondary antibody. Arrows and arrowheads indicate the cytoplasmic and nuclear dots, respectively. (D) Distribution of U14 and p53 in cytoplasmic and nuclear fractions. Mock (m)- and HST (i)-infected Molt-3 cells were harvested at 24 hpi and subjected to cytoplasmic and nuclear fractionation. Topoisomerase II α and α-tubulin were used as positive controls for the nuclear and cytoplasmic proteins, respectively.
    Fluorescein Isothiocyanate Fitc, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 93/100, based on 494 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Thermo Fisher annexin v fluorescein isothiocyanate
    (a) The control cells treated with 0.1% DMSO showed no induction of apoptosis. The cells treated with OSEO showed gradual increased in percentage of apoptotic cells at concentration of (b) 50 μg/ml (16%), (c) 100 μg/ml (38%), (d) 250 μg/ml (61%), (e) 500 μg/ml (84%). (f) The resveratrol 300 μg/ml treated cells showed about 54% of apoptotic cells population. OSEO: Ocimum sanctum essential oil; MCF-7: Michigan cancer foundation-7; Annexin V-FITC: <t>Annexin-V-fluorescein</t> <t>isothiocyanate;</t> PI: Propidium iodide
    Annexin V Fluorescein Isothiocyanate, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 801 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    Interaction of Candid 1 with VGCC subunits on the cell surface. (a) Cells treated with the indicated siRNAs and incubated with FITC-labeled Candid 1. Shown is a representative FACs plot and below the graph, the average MFI of the peaks averaged from 2

    Journal: Science translational medicine

    Article Title: siRNA screen for genes that affect Jun\u00edn virus entry uncovers voltage-gated calcium channels as a therapeutic target

    doi: 10.1126/scitranslmed.3006827

    Figure Lengend Snippet: Interaction of Candid 1 with VGCC subunits on the cell surface. (a) Cells treated with the indicated siRNAs and incubated with FITC-labeled Candid 1. Shown is a representative FACs plot and below the graph, the average MFI of the peaks averaged from 2

    Article Snippet: TfR1 levels were determined by FACS using human anti-CD71-PE or -FITC (Invitrogen).

    Techniques: Incubation, Labeling, FACS

    Rotavirus (RV) protein synthesis upon herpes simplex virus type-1 (HSV-1) amplicon vector transduction. ( A ) Schematic representation of the polycistronic gene expression cassettes packaged into herpesvirus amplicon particles. Herpes simplex virus type-1 amplicon particles deliver a DNA cassette comprising a single polycistronic messenger RNA containing the coding sequences of three RV capsid proteins VP2, VP6 and VP7 separated by internal ribosome entry sites (IRES) and controlled by the HSV-1 immediate early (IE) 4/5 promoter. The HSV-1 origin of DNA replication (oriS) and packaging/cleavage signal (pac) as well as the SV40 polyadenylation sequence (polyA) are indicated. The synthetic RV cassettes derived from the human Wa strain were adjusted to human codon usage (synthetic strain Wa, sWa). The RRV[VP7/6/2] was derived from the rhesus strain RRV and encodes, in addition to the RV proteins, for the enhanced green fluorescent protein (EGFP) as reporter gene. The sWa[VP2/6/7_V5] amplicon vector harbors a C-terminal V5 tag at VP7. The GFP amplicon vector encoding EGFP was used as control vector; ( B ) Rotavirus gene expression from codon optimized transgene cassettes. HepG2 cells were transduced with the indicated amplicon vectors (multiplicity of infection, MOI 2) or non-transduced (mock), and total cell lysates were harvested at 24 h post-transduction (hpt). Transgene expression was analyzed by Western blotting using the polyclonal anti-RV antibody to detect RV proteins (upper panel), the anti-V5 antibody for detection of the V5 tagged VP7 protein (middle panel), or the anti-actin and anti-GFP antibodies (lower panel). Detection of actin served as loading control. The predicted migration lengths of VP2 (104 kDa), VP6 (45 kDa), VP7 (37 kDa), VP7_V5 (42 kDa), actin (42 kDa) and GFP (27 kDa) are indicated on the left with arrows. The positions of the molecular weight markers are indicated on the right in kDa; ( C ) Analysis of protein synthesis in human and mouse cells. Human HepG2 or mouse Hepa 1-6 cells were transduced either with sWa[VP2/6/7], RRV[VP7/6/2] or GFP HSV-1 amplicon vectors (MOI 2) or non-transduced (mock) and total cell lysates were harvested 24 hpt. Transgene expression was analyzed using the polyclonal anti-RV antibody for detection of RV proteins (upper panels), the anti-actin antibody (middle panels) or the anti-GFP antibody (lower panels). Detection of actin served as loading control. The predicted migration lengths of the individual proteins are indicated on the left and the positions of the molecular weight markers are shown in kDa on the right; ( D ) Intracellular distribution of HSV-1 amplicon vector encoded RV proteins. Vero2-2 cells were transduced with the amplicon vector sWa[VP2/6/7_V5] at an MOI of 1 and analyzed by immune fluorescence 24 hpt with a Leica SP8 confocal laser scanning microscope. The RV proteins were stained using the following antibodies: anti-VP2 antibody directly labeled with Zenon Alexa-Fluor 594 (red), anti-VP6 and the anti-mouse Alexa-Fluor 633 (cyan), anti-V5 directly labeled with fluorescein isothiocyanate (FITC) for the V5-tagged VP7 (green). The endoplasmic reticulum (ER) was stained using the Alexa-Fluor 405 conjugated lectin concanavalin A (ConA) (grey).

    Journal: International Journal of Molecular Sciences

    Article Title: Transfer of Anti-Rotavirus Antibodies during Pregnancy and in Milk Following Maternal Vaccination with a Herpes Simplex Virus Type-1 Amplicon Vector

    doi: 10.3390/ijms18020431

    Figure Lengend Snippet: Rotavirus (RV) protein synthesis upon herpes simplex virus type-1 (HSV-1) amplicon vector transduction. ( A ) Schematic representation of the polycistronic gene expression cassettes packaged into herpesvirus amplicon particles. Herpes simplex virus type-1 amplicon particles deliver a DNA cassette comprising a single polycistronic messenger RNA containing the coding sequences of three RV capsid proteins VP2, VP6 and VP7 separated by internal ribosome entry sites (IRES) and controlled by the HSV-1 immediate early (IE) 4/5 promoter. The HSV-1 origin of DNA replication (oriS) and packaging/cleavage signal (pac) as well as the SV40 polyadenylation sequence (polyA) are indicated. The synthetic RV cassettes derived from the human Wa strain were adjusted to human codon usage (synthetic strain Wa, sWa). The RRV[VP7/6/2] was derived from the rhesus strain RRV and encodes, in addition to the RV proteins, for the enhanced green fluorescent protein (EGFP) as reporter gene. The sWa[VP2/6/7_V5] amplicon vector harbors a C-terminal V5 tag at VP7. The GFP amplicon vector encoding EGFP was used as control vector; ( B ) Rotavirus gene expression from codon optimized transgene cassettes. HepG2 cells were transduced with the indicated amplicon vectors (multiplicity of infection, MOI 2) or non-transduced (mock), and total cell lysates were harvested at 24 h post-transduction (hpt). Transgene expression was analyzed by Western blotting using the polyclonal anti-RV antibody to detect RV proteins (upper panel), the anti-V5 antibody for detection of the V5 tagged VP7 protein (middle panel), or the anti-actin and anti-GFP antibodies (lower panel). Detection of actin served as loading control. The predicted migration lengths of VP2 (104 kDa), VP6 (45 kDa), VP7 (37 kDa), VP7_V5 (42 kDa), actin (42 kDa) and GFP (27 kDa) are indicated on the left with arrows. The positions of the molecular weight markers are indicated on the right in kDa; ( C ) Analysis of protein synthesis in human and mouse cells. Human HepG2 or mouse Hepa 1-6 cells were transduced either with sWa[VP2/6/7], RRV[VP7/6/2] or GFP HSV-1 amplicon vectors (MOI 2) or non-transduced (mock) and total cell lysates were harvested 24 hpt. Transgene expression was analyzed using the polyclonal anti-RV antibody for detection of RV proteins (upper panels), the anti-actin antibody (middle panels) or the anti-GFP antibody (lower panels). Detection of actin served as loading control. The predicted migration lengths of the individual proteins are indicated on the left and the positions of the molecular weight markers are shown in kDa on the right; ( D ) Intracellular distribution of HSV-1 amplicon vector encoded RV proteins. Vero2-2 cells were transduced with the amplicon vector sWa[VP2/6/7_V5] at an MOI of 1 and analyzed by immune fluorescence 24 hpt with a Leica SP8 confocal laser scanning microscope. The RV proteins were stained using the following antibodies: anti-VP2 antibody directly labeled with Zenon Alexa-Fluor 594 (red), anti-VP6 and the anti-mouse Alexa-Fluor 633 (cyan), anti-V5 directly labeled with fluorescein isothiocyanate (FITC) for the V5-tagged VP7 (green). The endoplasmic reticulum (ER) was stained using the Alexa-Fluor 405 conjugated lectin concanavalin A (ConA) (grey).

    Article Snippet: Cells were incubated with the corresponding antibodies diluted in PBS-BSA: anti-VP2 antibody (1:200; provided by Didier Poncet, CNRS/INRA, Gif-sur-Yvette, France) directly labeled with Zenon Alexa-Fluor 594 according to the manufacturer’s instructions (Molecular Probes, Thermo Fisher Scientific), mouse monoclonal anti-VP6 (1:500; Novus Biologicals, Cambridge, UK) and the anti-mouse Alexa-Fluor 633 (1:500; Molecular Probes, Thermo Fisher Scientific), mouse monoclonal anti-V5 antibody directly labeled with fluorescein isothiocyanate (FITC) (1:500; Molecular Probes) for the V5-tagged VP7.

    Techniques: Amplification, Plasmid Preparation, Transduction, Expressing, Sequencing, Derivative Assay, Infection, Western Blot, Migration, Molecular Weight, Fluorescence, Laser-Scanning Microscopy, Staining, Labeling

    Activation of the m-calpain isoform in the ACT-treated cells. Western blot of the cytosolic fraction isolated from J774A.1 cells treated with ACT (35 nM) or from cells transfected with anti m-calpain siRNA and treated with ACT (35 nM) (A). Kinetics of the proteolytic cleavage of the FRET-based calpain specific substrate for ACT-treated cells (35 nM toxin), and cells in which m-calpain expression has been silenced by transfection with the corresponding siRNA, or mock cells transfected with scrambled control siRNA, or cells pre-incubated with SAJ6017 (78 nM), a specific m-calpain inhibitor. The figure is a representative experiment from three similar assays performed under identical conditions (B). Assay of calpain activity using FITC-labelled α-casein as substrate. FITC-α-casein hydrolysis by calpain is determined from in J774A.1 cell extracts incubated with FITC-labelled α-casein in the presence or absence of 1 mM CaCl 2 . Protease activity was quantified in 10% polyacrylamide gel by determining fluorescence intensity of the non-hydrolysed FITC-α-casein band. Non-digested FITC-α-casein and cell extracts without calcium were used as negative controls (C).

    Journal: PLoS ONE

    Article Title: Calpain-Mediated Processing of Adenylate Cyclase Toxin Generates a Cytosolic Soluble Catalytically Active N-Terminal Domain

    doi: 10.1371/journal.pone.0067648

    Figure Lengend Snippet: Activation of the m-calpain isoform in the ACT-treated cells. Western blot of the cytosolic fraction isolated from J774A.1 cells treated with ACT (35 nM) or from cells transfected with anti m-calpain siRNA and treated with ACT (35 nM) (A). Kinetics of the proteolytic cleavage of the FRET-based calpain specific substrate for ACT-treated cells (35 nM toxin), and cells in which m-calpain expression has been silenced by transfection with the corresponding siRNA, or mock cells transfected with scrambled control siRNA, or cells pre-incubated with SAJ6017 (78 nM), a specific m-calpain inhibitor. The figure is a representative experiment from three similar assays performed under identical conditions (B). Assay of calpain activity using FITC-labelled α-casein as substrate. FITC-α-casein hydrolysis by calpain is determined from in J774A.1 cell extracts incubated with FITC-labelled α-casein in the presence or absence of 1 mM CaCl 2 . Protease activity was quantified in 10% polyacrylamide gel by determining fluorescence intensity of the non-hydrolysed FITC-α-casein band. Non-digested FITC-α-casein and cell extracts without calcium were used as negative controls (C).

    Article Snippet: Briefly, α-casein (from bovine milk, Sigma-Aldrich) was labelled following the manufactureŕs protocol with FITC (isomer I, Sigma-Aldrich) at 1∶5 protein/fluorophore ratio.

    Techniques: Activation Assay, Activated Clotting Time Assay, Western Blot, Isolation, Transfection, Expressing, Incubation, Activity Assay, Fluorescence

    Immunohistochemistry for T. gondii detection using polyclonal IgY anti-STAg antibodies. Paraffin-embedded brain sections of mice chronically infected with ME-49 strain were incubated with IgY anti-STAg and rabbit IgG anti-IgY conjugated to fluorescein isothiocyanate (FITC). ( A and B ), segmented parasitophorus vacuoles were detected into host cell cytoplasm around the DAPI-stained nucleus (blue). ( C and D ), IgY anti-STAg antibodies strongly recognized antigens on outer walls of tissue cysts and free tachyzoites ( C , arrow).

    Journal: PLoS ONE

    Article Title: Production, Characterization and Applications for Toxoplasma gondii-Specific Polyclonal Chicken Egg Yolk Immunoglobulins

    doi: 10.1371/journal.pone.0040391

    Figure Lengend Snippet: Immunohistochemistry for T. gondii detection using polyclonal IgY anti-STAg antibodies. Paraffin-embedded brain sections of mice chronically infected with ME-49 strain were incubated with IgY anti-STAg and rabbit IgG anti-IgY conjugated to fluorescein isothiocyanate (FITC). ( A and B ), segmented parasitophorus vacuoles were detected into host cell cytoplasm around the DAPI-stained nucleus (blue). ( C and D ), IgY anti-STAg antibodies strongly recognized antigens on outer walls of tissue cysts and free tachyzoites ( C , arrow).

    Article Snippet: After washing, slides were incubated with rabbit anti-IgY labeled with fluorescein isothiocyanate (FITC, Sigma) diluted 1∶300 in addition to DAPI for 60 min at 37°C.

    Techniques: Immunohistochemistry, Mouse Assay, Infection, Incubation, Staining

    Immunocytochemistry for T. gondii using polyclonal IgY anti-STAg. HeLa cells infected with tachyzoites were fixed, permeabilized and incubated with IgY anti-STAg and rabbit IgG anti-IgY conjugated to fluorescein isothiocyanate (FITC). In fluorescence photomicrographs, or merged images with phase contrast. tachyzoites were detected into cell cytoplasm around the nucleus (blue, DAPI) with strong staining (arrows).

    Journal: PLoS ONE

    Article Title: Production, Characterization and Applications for Toxoplasma gondii-Specific Polyclonal Chicken Egg Yolk Immunoglobulins

    doi: 10.1371/journal.pone.0040391

    Figure Lengend Snippet: Immunocytochemistry for T. gondii using polyclonal IgY anti-STAg. HeLa cells infected with tachyzoites were fixed, permeabilized and incubated with IgY anti-STAg and rabbit IgG anti-IgY conjugated to fluorescein isothiocyanate (FITC). In fluorescence photomicrographs, or merged images with phase contrast. tachyzoites were detected into cell cytoplasm around the nucleus (blue, DAPI) with strong staining (arrows).

    Article Snippet: After washing, slides were incubated with rabbit anti-IgY labeled with fluorescein isothiocyanate (FITC, Sigma) diluted 1∶300 in addition to DAPI for 60 min at 37°C.

    Techniques: Immunocytochemistry, Infection, Incubation, Fluorescence, Staining

    Confocal microscopy imaging results. (A) Schematic representation of the fusion of cyclic arginine-glycine-aspartic (RGD) and octa-arginine (R8) peptide-modified ergosterol (ERG)-combined cisplatin (diamminedichloridoplatinum(II) [DDP]) targeting liposome (LIP, RGD/R8-DDP/ERG-LIP) with A549 tumor cells under different pH conditions. (B) Confocal imaging of co-localized RGD/R8-DDP/ERG-LIP, cell nuclei, and lysosomes under different pH conditions. The cell nuclei were stained with nuclear-specific dye 4′,6-diamidino-2-phenylindole (DAPI), RGD/R8-DDP/ERG-LIP was labeled with fluorescein isothiocyanate (FITC), and lysosomes were labeled with Lyso-Tracker Red (blue, green, and red fluorescence, respectively). DAPI = 4′,6- diamidino-2-phenylindole, DDP = cisplatin (diamminedichloridoplatinum(II)), ERG = ergosterol , FITC = fluorescein isothiocyanate, LIPs = liposomes.

    Journal: Medicine

    Article Title: Antilung cancer effect of ergosterol and cisplatin-loaded liposomes modified with cyclic arginine-glycine-aspartic acid and octa-arginine peptides

    doi: 10.1097/MD.0000000000011916

    Figure Lengend Snippet: Confocal microscopy imaging results. (A) Schematic representation of the fusion of cyclic arginine-glycine-aspartic (RGD) and octa-arginine (R8) peptide-modified ergosterol (ERG)-combined cisplatin (diamminedichloridoplatinum(II) [DDP]) targeting liposome (LIP, RGD/R8-DDP/ERG-LIP) with A549 tumor cells under different pH conditions. (B) Confocal imaging of co-localized RGD/R8-DDP/ERG-LIP, cell nuclei, and lysosomes under different pH conditions. The cell nuclei were stained with nuclear-specific dye 4′,6-diamidino-2-phenylindole (DAPI), RGD/R8-DDP/ERG-LIP was labeled with fluorescein isothiocyanate (FITC), and lysosomes were labeled with Lyso-Tracker Red (blue, green, and red fluorescence, respectively). DAPI = 4′,6- diamidino-2-phenylindole, DDP = cisplatin (diamminedichloridoplatinum(II)), ERG = ergosterol , FITC = fluorescein isothiocyanate, LIPs = liposomes.

    Article Snippet: Furthermore, ERG, DDP, and fluorescein isothiocyanate (FITC) were purchased from Sigma-Aldrich Corporation, St. Louis.

    Techniques: Confocal Microscopy, Imaging, Modification, Staining, Labeling, Fluorescence

    Grape seed proanthocyanidin extract (GSPE) effect on physiological parameters. Wistar rats that received GSPE treatments are indicated as white columns. Control animals are indicated as black columns. ( A ) rats received a chronic treatment with 0.5 g GSPE /kg body weight (BW). Fluorescein isothiocyanate (FITC) was orally forced and FITC in different gastrointestinal localizations was measured after death; ( B ) rats received an acute treatment of 1 g GSPE/kg BW. After death, hypothalamic CART expression was measured. n = 6 per group; * p

    Journal: Nutrients

    Article Title: Defining Conditions for Optimal Inhibition of Food Intake in Rats by a Grape-Seed Derived Proanthocyanidin Extract

    doi: 10.3390/nu8100652

    Figure Lengend Snippet: Grape seed proanthocyanidin extract (GSPE) effect on physiological parameters. Wistar rats that received GSPE treatments are indicated as white columns. Control animals are indicated as black columns. ( A ) rats received a chronic treatment with 0.5 g GSPE /kg body weight (BW). Fluorescein isothiocyanate (FITC) was orally forced and FITC in different gastrointestinal localizations was measured after death; ( B ) rats received an acute treatment of 1 g GSPE/kg BW. After death, hypothalamic CART expression was measured. n = 6 per group; * p

    Article Snippet: Gallic acid, exendin 9-39 (Ex9) and 70 kDa fluorescein isothiocyanate (FITC)-dextran were obtained from Sigma (St. Louis, MO, USA).

    Techniques: Expressing

    Microbubble (MB) binding to receptor expression. Correlation of MB EGFR (A), MB CD147 (B), and MB DUAL (C) binding to receptor expression was assessed in 4 head and neck cancer cell lines (SCC-1, SCC-5, FaDu, and Cal27), 2 head and neck cancer CD147 knock-down cell lines (SCC-1/SiE-711 and SCC-5/SiE-711), and normal dermal fibroblasts (NDFs). FITC indicates fluorescein isothiocyanate. MB IgG , MB EGFR , MB CD147 , and MB DUAL indicate IgG-, anti-EGFR–, and anti-CD147–targeted MBs and dual-conjugated MBs, respectively.

    Journal: Archives of otolaryngology--head & neck surgery

    Article Title: Molecular Targeting of Ultrasonographic Contrast Agent for Detection of Head and Neck Squamous Cell Carcinoma

    doi: 10.1001/archoto.2012.1081

    Figure Lengend Snippet: Microbubble (MB) binding to receptor expression. Correlation of MB EGFR (A), MB CD147 (B), and MB DUAL (C) binding to receptor expression was assessed in 4 head and neck cancer cell lines (SCC-1, SCC-5, FaDu, and Cal27), 2 head and neck cancer CD147 knock-down cell lines (SCC-1/SiE-711 and SCC-5/SiE-711), and normal dermal fibroblasts (NDFs). FITC indicates fluorescein isothiocyanate. MB IgG , MB EGFR , MB CD147 , and MB DUAL indicate IgG-, anti-EGFR–, and anti-CD147–targeted MBs and dual-conjugated MBs, respectively.

    Article Snippet: All human cancer cells were aliquoted (100 000 cells per tube) and stained with fluorescein isothiocyanate conjugated to primary antibodies against either human IgG control, EGFR, or CD147 (BD Pharmingen).

    Techniques: Binding Assay, Expressing

    Bar graft of flow cytometry analysis. Four head and neck cancer cell lines (SCC-1, SCC-5, FaDu, and Cal27), 2 CD147 knock-down cell lines (SCC-1/SiE-711 and SCC-5/SiE-711), and normal dermal fibroblasts (NDFs) were analyzed by flow cytometry to assess cell surface expression of epidermal growth factor receptor (EGFR) and extracellular matrix-metalloproteinase inhibitor (CD147). FITC indicates fluorescein isothiocyanate. Error bars indicate SE. ** P

    Journal: Archives of otolaryngology--head & neck surgery

    Article Title: Molecular Targeting of Ultrasonographic Contrast Agent for Detection of Head and Neck Squamous Cell Carcinoma

    doi: 10.1001/archoto.2012.1081

    Figure Lengend Snippet: Bar graft of flow cytometry analysis. Four head and neck cancer cell lines (SCC-1, SCC-5, FaDu, and Cal27), 2 CD147 knock-down cell lines (SCC-1/SiE-711 and SCC-5/SiE-711), and normal dermal fibroblasts (NDFs) were analyzed by flow cytometry to assess cell surface expression of epidermal growth factor receptor (EGFR) and extracellular matrix-metalloproteinase inhibitor (CD147). FITC indicates fluorescein isothiocyanate. Error bars indicate SE. ** P

    Article Snippet: All human cancer cells were aliquoted (100 000 cells per tube) and stained with fluorescein isothiocyanate conjugated to primary antibodies against either human IgG control, EGFR, or CD147 (BD Pharmingen).

    Techniques: Flow Cytometry, Cytometry, Expressing

    Percentage of Th1, Th2, and Th0 cells in nonpregnant women, normal pregnant women, and preeclamptic patients. Peripheral blood mononuclear cells were stained with Peridinin chlorophyll protein (PerCP)-conjugated CD4 monoclonal antibody, fluorescein-isothiocyanate (FITC)-conjugated anti IFN-γ monoclonal antibody and PE-conjugated anti-interleukin (IL)-4 monoclonal antibody, as described in Materials and Methods. A gate was set on the CD4 + lymphocytes by characteristic forward and side scatter parameters (R1 and R2). The FITC (anti-interferon (IFN)-γ) and PE (anti-IL-4) fluorescence data were analysed. Isotype muched fluorochrome-conjugated IgG1 and IgG2a were used as control. The horizontal bar shows FITC fluorescence and the vertical bar shows PE fluorescence. Fluorescence intensity is determined on a logarithmic scale.

    Journal: Clinical and Experimental Immunology

    Article Title: Quantitative analysis of peripheral blood Th0, Th1, Th2 and the Th1:Th2 cell ratio during normal human pregnancy and preeclampsia

    doi: 10.1046/j.1365-2249.1999.00997.x

    Figure Lengend Snippet: Percentage of Th1, Th2, and Th0 cells in nonpregnant women, normal pregnant women, and preeclamptic patients. Peripheral blood mononuclear cells were stained with Peridinin chlorophyll protein (PerCP)-conjugated CD4 monoclonal antibody, fluorescein-isothiocyanate (FITC)-conjugated anti IFN-γ monoclonal antibody and PE-conjugated anti-interleukin (IL)-4 monoclonal antibody, as described in Materials and Methods. A gate was set on the CD4 + lymphocytes by characteristic forward and side scatter parameters (R1 and R2). The FITC (anti-interferon (IFN)-γ) and PE (anti-IL-4) fluorescence data were analysed. Isotype muched fluorochrome-conjugated IgG1 and IgG2a were used as control. The horizontal bar shows FITC fluorescence and the vertical bar shows PE fluorescence. Fluorescence intensity is determined on a logarithmic scale.

    Article Snippet: These permeabilized cells were stained with fluorescein-isothiocyanate (FITC) labelled anti-human IFN-γ monoclonal antibody (Becton Dickinson), and PE-labelled anti-human IL-4 monoclonal antibody (Becton Dickinson).

    Techniques: Staining, Fluorescence

    UDP inhibits C5a-stimulated macropinocytosis. BMDM were stimulated with or without C5a (0, 0.3, 0.75, and 2.5 n m ) in the presence or absence of UDP (0, 0.5, and 2.5 μ m ) in assays of macropinocytosis. Uptake of extracellular fluorescein isothiocyanate-dextran

    Journal:

    Article Title: Signaling and Cross-talk by C5a and UDP in Macrophages Selectively Use PLC?3 to Regulate Intracellular Free Calcium *Signaling and Cross-talk by C5a and UDP in Macrophages Selectively Use PLC?3 to Regulate Intracellular Free Calcium * S⃞

    doi: 10.1074/jbc.M800907200

    Figure Lengend Snippet: UDP inhibits C5a-stimulated macropinocytosis. BMDM were stimulated with or without C5a (0, 0.3, 0.75, and 2.5 n m ) in the presence or absence of UDP (0, 0.5, and 2.5 μ m ) in assays of macropinocytosis. Uptake of extracellular fluorescein isothiocyanate-dextran

    Article Snippet: Reagents —UDP, UTP, LPA, platelet activating factor (PAF), human C5a, and fluorescein isothiocyanate-dextran were from Sigma.

    Techniques:

    DSPS alters cell cycle progression and promotes apoptosis. ( a ) MDA‐MB‐231 cells treated with various concentrations of DSPS were stained with PI, and the cells were analyzed by flow cytometry to determine cell cycle progression. ( b ) Quantification of the percentage of cells in G1, S, and G2/M phases of cells treated as in panel A. 0 mg/mL, 0.5 mg/mL, 5 mg/mL. ( c–f ) MDA‐MB‐231 cells treated with various concentrations of DSPS and stained with Annexin V fluorescein isothiocyanate (FITC)/propidium iodide ( c ); the percentage of total apoptotic cells ( d ), cells in early apoptosis ( e ), and cells in late apoptosis ( f ) were quantified.

    Journal: Thoracic Cancer

    Article Title: Exopolysaccharides from a Codonopsis pilosula endophyte activate macrophages and inhibit cancer cell proliferation and migration

    doi: 10.1111/1759-7714.12630

    Figure Lengend Snippet: DSPS alters cell cycle progression and promotes apoptosis. ( a ) MDA‐MB‐231 cells treated with various concentrations of DSPS were stained with PI, and the cells were analyzed by flow cytometry to determine cell cycle progression. ( b ) Quantification of the percentage of cells in G1, S, and G2/M phases of cells treated as in panel A. 0 mg/mL, 0.5 mg/mL, 5 mg/mL. ( c–f ) MDA‐MB‐231 cells treated with various concentrations of DSPS and stained with Annexin V fluorescein isothiocyanate (FITC)/propidium iodide ( c ); the percentage of total apoptotic cells ( d ), cells in early apoptosis ( e ), and cells in late apoptosis ( f ) were quantified.

    Article Snippet: Fluorescein isothiocyanate (FITC)‐conjugated phalloidin was obtained from ThermoFisher Scientific (Waltham, MA, USA).

    Techniques: Multiple Displacement Amplification, Staining, Flow Cytometry, Cytometry

    DSPS affects cell migration. ( a ) Transwell migration assay showing the effect of DSPS treatment on RAW264.7 cell migration. ( b ) Quantification of cell migration; cells were treated as in panel ( a ). ( c ) Immunofluorescence staining of actin (fluorescein isothiocyanate‐conjugated phalloidin) and nuclei (4′, 6‐diamidino‐2‐phenylindole) in RAW264.7 cells. Scale bars, 5 μm. ( d ) Quantification of filopodia presented in panel ( c ). ( e ) Representative wound healing assay analyzing the effects of DSPS on the migration of BT549 and MDA‐MB‐231 breast cancer cells. ( f ) Quantification of migrated cells treated as in panel ( e ).

    Journal: Thoracic Cancer

    Article Title: Exopolysaccharides from a Codonopsis pilosula endophyte activate macrophages and inhibit cancer cell proliferation and migration

    doi: 10.1111/1759-7714.12630

    Figure Lengend Snippet: DSPS affects cell migration. ( a ) Transwell migration assay showing the effect of DSPS treatment on RAW264.7 cell migration. ( b ) Quantification of cell migration; cells were treated as in panel ( a ). ( c ) Immunofluorescence staining of actin (fluorescein isothiocyanate‐conjugated phalloidin) and nuclei (4′, 6‐diamidino‐2‐phenylindole) in RAW264.7 cells. Scale bars, 5 μm. ( d ) Quantification of filopodia presented in panel ( c ). ( e ) Representative wound healing assay analyzing the effects of DSPS on the migration of BT549 and MDA‐MB‐231 breast cancer cells. ( f ) Quantification of migrated cells treated as in panel ( e ).

    Article Snippet: Fluorescein isothiocyanate (FITC)‐conjugated phalloidin was obtained from ThermoFisher Scientific (Waltham, MA, USA).

    Techniques: Migration, Transwell Migration Assay, Immunofluorescence, Staining, Wound Healing Assay, Multiple Displacement Amplification

    3-MA significantly increased the apoptotic rate under hypoxic conditions. Apoptosis was detected by a Annexin V-fluorescein isothiocyanate assay. (A) Flow cytometry analysis of apoptosis. Q2 represented the early apoptotic cells. Q4 represented the late apoptotic cells. Apoptotic cells from each group were detected by Annexin V and propidium iodide. (B) The proportion of apoptotic cells from each group was quantified. Data are presented as the mean ± standard deviation. *P

    Journal: Molecular Medicine Reports

    Article Title: Inhibition of autophagy enhanced cobalt chloride-induced apoptosis in rat alveolar type II epithelial cells

    doi: 10.3892/mmr.2018.9209

    Figure Lengend Snippet: 3-MA significantly increased the apoptotic rate under hypoxic conditions. Apoptosis was detected by a Annexin V-fluorescein isothiocyanate assay. (A) Flow cytometry analysis of apoptosis. Q2 represented the early apoptotic cells. Q4 represented the late apoptotic cells. Apoptotic cells from each group were detected by Annexin V and propidium iodide. (B) The proportion of apoptotic cells from each group was quantified. Data are presented as the mean ± standard deviation. *P

    Article Snippet: The Annexin V-fluorescein isothiocyanate (FITC)/PI apoptosis detection kit was purchased from BD Biosciences (San Jose, CA, USA).

    Techniques: Flow Cytometry, Cytometry, Standard Deviation

    Hyperthermia increases GEM-induced cell apoptosis. (A) SW1990 cells were treated with hyperthermia at 42°C for 90 min, and then were treated with 5 µM GEM for 24 h. Cell apoptosis was assessed using an Annexin V-fluorescein isothiocyanate/propidium iodide assay. (B) Morphological changes of SW1990 cells were observed under a transmission electron microscope (magnification, ×6,000), including the following groups: (a) Normal cells, (b) cells treated with hyperthermia alone, (c) cells treated with GEM alone and (d) cells treated with hyperthermia combined with GEM. (C) Cell cycle distributions of PC cells were detected using flow cytometry. (D) Protein expression of the apoptosis-associated proteins, B-cell lymphoma 2, Bcl-2-associated X protein, cleaved caspase-3 and cleaved caspase-9 were evaluated using western blot analysis using respective antibodies. β-actin was detected as internal reference. *P

    Journal: Oncology Letters

    Article Title: Hyperthermia enhances the sensitivity of pancreatic cancer SW1990 cells to gemcitabine through ROS/JNK signaling

    doi: 10.3892/ol.2018.9455

    Figure Lengend Snippet: Hyperthermia increases GEM-induced cell apoptosis. (A) SW1990 cells were treated with hyperthermia at 42°C for 90 min, and then were treated with 5 µM GEM for 24 h. Cell apoptosis was assessed using an Annexin V-fluorescein isothiocyanate/propidium iodide assay. (B) Morphological changes of SW1990 cells were observed under a transmission electron microscope (magnification, ×6,000), including the following groups: (a) Normal cells, (b) cells treated with hyperthermia alone, (c) cells treated with GEM alone and (d) cells treated with hyperthermia combined with GEM. (C) Cell cycle distributions of PC cells were detected using flow cytometry. (D) Protein expression of the apoptosis-associated proteins, B-cell lymphoma 2, Bcl-2-associated X protein, cleaved caspase-3 and cleaved caspase-9 were evaluated using western blot analysis using respective antibodies. β-actin was detected as internal reference. *P

    Article Snippet: Apoptosis was analyzed using an Annexin V-fluorescein isothiocyanate (FITC)/propidium iodide (PI) detection kit (cat. no. 556547; BD Biosciences, Franklin Lakes, NJ, USA), according to the manufacturer's protocol.

    Techniques: Transmission Assay, Microscopy, Flow Cytometry, Cytometry, Expressing, Western Blot

    Effect of different concentrations of udenafil for 24 h on apoptosis in VEC and VSMC. Distributions of cells treated with different concentrations of udenafil displayed as dot plots: Viable cells (fluorescein isothiocyanate (FITC˜)/propidium iodide (PĨ)), apoptotic cells (FITC+/PĨ), secondary necrotic cells (FITC+/PI+). (a and f) Cells incubated with 0 μmol/l udenafil. (b and g) Cells incubated with 1 μmol/l udenafil. (c and h) Cells incubated with 10 μmol/l udenafil. (d and i) Cells incubated with 100 μmol/l udenafil. (e and j) Cells incubated with 1 mmol/l udenafil. A minimum of 10,000 events was counted per sample

    Journal: Indian Journal of Pharmacology

    Article Title: Concentration-dependent differential effects of udenafil on viability, proliferation, and apoptosis in vascular endothelial and smooth muscle cells

    doi: 10.4103/0253-7613.132161

    Figure Lengend Snippet: Effect of different concentrations of udenafil for 24 h on apoptosis in VEC and VSMC. Distributions of cells treated with different concentrations of udenafil displayed as dot plots: Viable cells (fluorescein isothiocyanate (FITC˜)/propidium iodide (PĨ)), apoptotic cells (FITC+/PĨ), secondary necrotic cells (FITC+/PI+). (a and f) Cells incubated with 0 μmol/l udenafil. (b and g) Cells incubated with 1 μmol/l udenafil. (c and h) Cells incubated with 10 μmol/l udenafil. (d and i) Cells incubated with 100 μmol/l udenafil. (e and j) Cells incubated with 1 mmol/l udenafil. A minimum of 10,000 events was counted per sample

    Article Snippet: After washing with phosphate-buffered saline (PBS), the cells were cultured with medium containing different concentrations of udenafil for 24 h. They were harvested with trypsin-ethylenediaminetetraacetic acid (EDTA), washed twice with cold PBS, and resuspended in 1 × binding buffer containing 5 μl annexin V-fluorescein isothiocyanate (FITC) and 5 μl propidium iodide solution (BD Biosciences, San Diego, CA, USA), and incubated for 15 min at room temperature.

    Techniques: Incubation

    Involvement of caspases in ferrugone and DMI-induced cell death in human ovarian cancer cells. A2780 cells were pretreated with broad caspase inhibitor z-VAD-fmk (50 µM) for 2 h, and then treated with ferrugone ( A ) and DMI ( B ) (0.5 µM) for 48 h. Annexin/PI V-FITC staining assay was performed to determine apoptosis. The graph indicates the percentages of annexin V-positive apoptotic cells in the right quadrants of flow cytometry graphs. The data are representative of three independent experiments. Student’s t -test (two-tailed) was applied to evaluate the significance. # p

    Journal: Molecules

    Article Title: Isoflavones Isolated from the Seeds of Millettia ferruginea Induced Apoptotic Cell Death in Human Ovarian Cancer Cells

    doi: 10.3390/molecules25010207

    Figure Lengend Snippet: Involvement of caspases in ferrugone and DMI-induced cell death in human ovarian cancer cells. A2780 cells were pretreated with broad caspase inhibitor z-VAD-fmk (50 µM) for 2 h, and then treated with ferrugone ( A ) and DMI ( B ) (0.5 µM) for 48 h. Annexin/PI V-FITC staining assay was performed to determine apoptosis. The graph indicates the percentages of annexin V-positive apoptotic cells in the right quadrants of flow cytometry graphs. The data are representative of three independent experiments. Student’s t -test (two-tailed) was applied to evaluate the significance. # p

    Article Snippet: Phenylmethylsulfonylfluoride (PMSF) and annexin V-fluorescein isothiocyanate (FITC) were purchased from BD Biosciences (San Jose, CA, USA).

    Techniques: Staining, Flow Cytometry, Two Tailed Test

    Effect of ferrugone and DMI on apoptotic cell death in human ovarian cancer cells. A2780 cells were treated with ferrugone ( A ) and DMI ( B ) (0.25, 0.5, and 1 µM) for 48 h and co-stained with PI and Annexin V-FITC (fluorescein isothiocyanate). The translocation of phosphatidyl serine was detected by flow cytometry. The graph indicates the percentages of annexin V-positive apoptotic cells in the right quadrants of flow cytometry graphs. The data are representative of three independent experiments. Data were analyzed using one-way ANOVA followed by Dunnett’s multiple comparison test. * p

    Journal: Molecules

    Article Title: Isoflavones Isolated from the Seeds of Millettia ferruginea Induced Apoptotic Cell Death in Human Ovarian Cancer Cells

    doi: 10.3390/molecules25010207

    Figure Lengend Snippet: Effect of ferrugone and DMI on apoptotic cell death in human ovarian cancer cells. A2780 cells were treated with ferrugone ( A ) and DMI ( B ) (0.25, 0.5, and 1 µM) for 48 h and co-stained with PI and Annexin V-FITC (fluorescein isothiocyanate). The translocation of phosphatidyl serine was detected by flow cytometry. The graph indicates the percentages of annexin V-positive apoptotic cells in the right quadrants of flow cytometry graphs. The data are representative of three independent experiments. Data were analyzed using one-way ANOVA followed by Dunnett’s multiple comparison test. * p

    Article Snippet: Phenylmethylsulfonylfluoride (PMSF) and annexin V-fluorescein isothiocyanate (FITC) were purchased from BD Biosciences (San Jose, CA, USA).

    Techniques: Staining, Translocation Assay, Flow Cytometry

    Immunofluorescence analysis of virus maturation. HFF were infected with RV wt AD169 or RV del UL35 at an MOI of 0.2. Cells were fixed at indicated time points (24, 72, and 120 hpi) and stained with monoclonal antibodies directed against ppUL82 (CMV355), ppUL83 (28-77), ppUL99 (41-18), and gpUL55 (27-287), as indicated at the top. Goat anti-mouse secondary antibody labeled with fluorescein isothiocyanate was used for detection. The nuclei were stained with DAPI. See the text for details.

    Journal: Journal of Virology

    Article Title: Human Cytomegalovirus Tegument Protein ppUL35 Is Important for Viral Replication and Particle Formation

    doi: 10.1128/JVI.79.5.3084-3096.2005

    Figure Lengend Snippet: Immunofluorescence analysis of virus maturation. HFF were infected with RV wt AD169 or RV del UL35 at an MOI of 0.2. Cells were fixed at indicated time points (24, 72, and 120 hpi) and stained with monoclonal antibodies directed against ppUL82 (CMV355), ppUL83 (28-77), ppUL99 (41-18), and gpUL55 (27-287), as indicated at the top. Goat anti-mouse secondary antibody labeled with fluorescein isothiocyanate was used for detection. The nuclei were stained with DAPI. See the text for details.

    Article Snippet: Subsequently, murine monoclonal antibodies directed against ppUL69 (antibody 69-66), ppUL82 (pp71) (antibody CMV355), ppUL83 (pp65) (antibody 28-77), ppUL99 (pp28) (antibody 41-18), or gpUL55 (gB) (antibody 27-287) were added as undiluted cell culture supernatants and incubated at 4°C for 2 h. After being washed three times for 5 min with PBS, the coverslips were incubated at 4°C for 2 h with anti-mouse goat serum labeled with fluorescein isothiocyanate as a secondary antibody (DAKO), which was used at a 1:50 dilution.

    Techniques: Immunofluorescence, Infection, Staining, Labeling

    Cholangiocarcinoma QBC939 cells apoptosis induced by solamargine. (A) Apoptosis of QBC939 cells induced by treatment with different concentrations of solamargine for 24 h, stained using Annexin V-FITC/PI and determined using flow cytometry. (B) Statistical data of QBC939 cell apoptosis. Apoptosis rate contains the early apoptosis rate and late apoptosis rate. *P

    Journal: Oncology Letters

    Article Title: Solamargine derived from Solanum nigrum induces apoptosis of human cholangiocarcinoma QBC939 cells

    doi: 10.3892/ol.2018.8171

    Figure Lengend Snippet: Cholangiocarcinoma QBC939 cells apoptosis induced by solamargine. (A) Apoptosis of QBC939 cells induced by treatment with different concentrations of solamargine for 24 h, stained using Annexin V-FITC/PI and determined using flow cytometry. (B) Statistical data of QBC939 cell apoptosis. Apoptosis rate contains the early apoptosis rate and late apoptosis rate. *P

    Article Snippet: Cells in each group (0, 2, 4, 6, 8 and 10 µM) were stained with 5 µl propidium iodide (PI) and 5 µl Annexin V-FITC, according to the Annexin V-fluorescein isothiocyanate (FITC) apoptosis detection kit (BD Biosciences, Franklin Lakes, NJ, USA) protocol and incubated for 15 min at room temperature in the dark.

    Techniques: Staining, Flow Cytometry, Cytometry

    Oral squamous cell carcinoma (OSCC) cells exhibit differential responses to irradiation (IR). ( A, B ) The effects of IR on the ability of OSCC cell lines to form colonies were determined on Day 14 after IR. The representative results and quantitative data from three independent experiments are shown in ( A ) and ( B ), respectively. ( C ) The effects of IR on OSCC cell viability were determined on Day 3 after IR. ( D ) The effect of IR on poly ADP-ribose polymerase (PARP) cleavage were determined in OSCC cell lines on Day 3 after IR. ( E, F ) The effects of IR on apoptosis were determined in OSCC cell lines using Fluorescein isothiocyanate (FITC) Annexin V Apoptosis Detection Kit with PI on Day 3 after IR. A representative result and quantitative data from three independent experiments are shown in ( E ) and ( F ), respectively. * p

    Journal: Cells

    Article Title: Circumventing AKT-Associated Radioresistance in Oral Cancer by Novel Nanoparticle-Encapsulated Capivasertib

    doi: 10.3390/cells9030533

    Figure Lengend Snippet: Oral squamous cell carcinoma (OSCC) cells exhibit differential responses to irradiation (IR). ( A, B ) The effects of IR on the ability of OSCC cell lines to form colonies were determined on Day 14 after IR. The representative results and quantitative data from three independent experiments are shown in ( A ) and ( B ), respectively. ( C ) The effects of IR on OSCC cell viability were determined on Day 3 after IR. ( D ) The effect of IR on poly ADP-ribose polymerase (PARP) cleavage were determined in OSCC cell lines on Day 3 after IR. ( E, F ) The effects of IR on apoptosis were determined in OSCC cell lines using Fluorescein isothiocyanate (FITC) Annexin V Apoptosis Detection Kit with PI on Day 3 after IR. A representative result and quantitative data from three independent experiments are shown in ( E ) and ( F ), respectively. * p

    Article Snippet: For cultured cells, apoptosis was assessed by flow cytometry using Fluorescein isothiocyanate (FITC) Annexin V Apoptosis Detection Kit (BD Biosciences, San Jose, CA, USA) with Propidium Iodide (PI).

    Techniques: Irradiation

    Interferon (IFN)-γ in combination with TNF-α induces cell death in mouse lung carcinoma cells. L929 fibroblasts and Lewis lung carcinoma (LLC) cancer cells were cultured for 24 h before starting treatment with recombinant cytokines. Both L929 and LLC cells were (A,B) left untreated for 48 h and used as negative controls or (C) L929 cells were treated with 50 ng/mL TNF-α for 48 h and used as a positive control. LLC cell viability was retained after treatment with (D) 50 ng/mL TNF-α or (E) 100 ng/mL IFN-γ and after (F) simultaneous treatment with both 50 ng/mL TNF-α and 100 ng/mL IFN-γ for 24 h. Cell death was induced in LLC cells after (G) simultaneous treatment with 100 ng/mL of IFN-γ in combination with 50 ng/mL of TNF-α for 48 h and after (H) LLC pretreatment with 100 ng/mL of IFN-γ for 24 h followed by treatment with TNF-α for 24 h. The resulting cell monolayers were analyzed using bright-field microscopy (scale bar, 50 µM). Cell death was determined using flow cytometry analysis after cell staining with annexin V-FITC (depicted on the x -axis) and propidium iodide (PI, depicted on the y -axis). Annexin V-positive/PI-negative cells were regarded as apoptotic, whereas annexin V-positive/PI-positive cells were regarded as necrotic. Experiment was performed in duplicates and repeated two times with standard error not exceeding 10% between independent experiments. One representative experiment is shown, where the percentages of the four distinct cell populations are averages of duplicates with SEM

    Journal: Frontiers in Immunology

    Article Title: Generation and Functional In Vitro Analysis of Semliki Forest Virus Vectors Encoding TNF-α and IFN-γ

    doi: 10.3389/fimmu.2017.01667

    Figure Lengend Snippet: Interferon (IFN)-γ in combination with TNF-α induces cell death in mouse lung carcinoma cells. L929 fibroblasts and Lewis lung carcinoma (LLC) cancer cells were cultured for 24 h before starting treatment with recombinant cytokines. Both L929 and LLC cells were (A,B) left untreated for 48 h and used as negative controls or (C) L929 cells were treated with 50 ng/mL TNF-α for 48 h and used as a positive control. LLC cell viability was retained after treatment with (D) 50 ng/mL TNF-α or (E) 100 ng/mL IFN-γ and after (F) simultaneous treatment with both 50 ng/mL TNF-α and 100 ng/mL IFN-γ for 24 h. Cell death was induced in LLC cells after (G) simultaneous treatment with 100 ng/mL of IFN-γ in combination with 50 ng/mL of TNF-α for 48 h and after (H) LLC pretreatment with 100 ng/mL of IFN-γ for 24 h followed by treatment with TNF-α for 24 h. The resulting cell monolayers were analyzed using bright-field microscopy (scale bar, 50 µM). Cell death was determined using flow cytometry analysis after cell staining with annexin V-FITC (depicted on the x -axis) and propidium iodide (PI, depicted on the y -axis). Annexin V-positive/PI-negative cells were regarded as apoptotic, whereas annexin V-positive/PI-positive cells were regarded as necrotic. Experiment was performed in duplicates and repeated two times with standard error not exceeding 10% between independent experiments. One representative experiment is shown, where the percentages of the four distinct cell populations are averages of duplicates with SEM

    Article Snippet: To evaluate cell viability BHK-21 and LLC cells were labeled 48 h post-infection with 1 µg/mL of fluorescein isothiocyanate (FITC)-conjugated annexin V (annexin V-FITC from Apoptosis Detection Kit; Cat. No. 556547; BD Biosciences) for 15 min followed by staining with 3 µM of DAPI (Cat. No. 422801; BioLegend, San Diego, CA, USA) for 5 min.

    Techniques: Cell Culture, Recombinant, Positive Control, Microscopy, Flow Cytometry, Cytometry, Staining

    Vector-derived TNF-α induces cell death in L929 fibroblasts. L929 cells were cultured for 24 h until they reached 90% confluency. The cells were then (A) left untreated for 24 h and used as a negative control or (B) treated with 1 µM staurosporine for 24 h at 37°C and used as a positive control. The remaining cells were treated with rTNF-α or vdTNF-α for 24 h at the following concentrations: (C,D) 2.2 ng/mL, (E,F) 6.7 ng/mL, (G,H) 20 ng/mL or (I,J) 60 ng/mL. The resulting cell monolayers were analyzed using bright-field microscopy (scale bar, 50 µM). Cell death was determined using flow cytometry analysis after cell staining with annexin V-FITC (depicted on the x -axis) and propidium iodide (PI, depicted on the y -axis). Annexin V-positive/PI-negative cells were regarded as apoptotic, whereas annexin V-positive/PI-positive cells were regarded as necrotic. Two independent experiments were performed in duplicates with standard error not exceeding 10% between independent experiments. Data from one representative experiment are shown, where the percentages of the four distinct cell populations represent the averages of duplicates with SEM

    Journal: Frontiers in Immunology

    Article Title: Generation and Functional In Vitro Analysis of Semliki Forest Virus Vectors Encoding TNF-α and IFN-γ

    doi: 10.3389/fimmu.2017.01667

    Figure Lengend Snippet: Vector-derived TNF-α induces cell death in L929 fibroblasts. L929 cells were cultured for 24 h until they reached 90% confluency. The cells were then (A) left untreated for 24 h and used as a negative control or (B) treated with 1 µM staurosporine for 24 h at 37°C and used as a positive control. The remaining cells were treated with rTNF-α or vdTNF-α for 24 h at the following concentrations: (C,D) 2.2 ng/mL, (E,F) 6.7 ng/mL, (G,H) 20 ng/mL or (I,J) 60 ng/mL. The resulting cell monolayers were analyzed using bright-field microscopy (scale bar, 50 µM). Cell death was determined using flow cytometry analysis after cell staining with annexin V-FITC (depicted on the x -axis) and propidium iodide (PI, depicted on the y -axis). Annexin V-positive/PI-negative cells were regarded as apoptotic, whereas annexin V-positive/PI-positive cells were regarded as necrotic. Two independent experiments were performed in duplicates with standard error not exceeding 10% between independent experiments. Data from one representative experiment are shown, where the percentages of the four distinct cell populations represent the averages of duplicates with SEM

    Article Snippet: To evaluate cell viability BHK-21 and LLC cells were labeled 48 h post-infection with 1 µg/mL of fluorescein isothiocyanate (FITC)-conjugated annexin V (annexin V-FITC from Apoptosis Detection Kit; Cat. No. 556547; BD Biosciences) for 15 min followed by staining with 3 µM of DAPI (Cat. No. 422801; BioLegend, San Diego, CA, USA) for 5 min.

    Techniques: Plasmid Preparation, Derivative Assay, Cell Culture, Negative Control, Positive Control, Microscopy, Flow Cytometry, Cytometry, Staining

    Macrophages remain viable, whereas cancer cells undergo cell death after challenge with Semliki Forest virus (SFV). J774A.1 mouse macrophages and Lewis lung carcinoma (LLC) carcinoma cells were infected with SFV-DsRed particles at MOI = 10 [determined using baby hamster kidney (BHK-21) cells]. DsRed expression (first column, x -axis) was evaluated at 48 h post-infection using flow cytometry and fluorescence microscopy (scale bar 100 µM). Cell morphology was observed using bright-field microscopy (scale bar 100 µM) and cell death was quantified 48 h post-infection using flow cytometry analysis after cell staining with annexin V-FITC (second column, x -axis) and DAPI (second column, y -axis). Annexin V-positive/DAPI-negative cells were regarded as early apoptotic, whereas annexin V-positive/PI-positive cells were regarded as late apoptotic and necrotic. (A) Non-infected J774A.1 macrophages were used as a negative control. (B) J774A.1 macrophages were resistant to SFV infection, stayed viable but changed their morphology 48 h after challenge with SFV. (C) Non-infected LLC cells were used as a negative control. (D) LLC cells were susceptible to SFV infection and underwent cell death. DsRed expression was observed using microscopy in both apoptotic (white arrows) and viable (yellow arrows) cells, whereas some apoptotic cells lacked DsRed expression (red arrows). (E) The infected DsRed-positive cells were both viable and undergoing cell death. The average of duplicates with SEM

    Journal: Frontiers in Immunology

    Article Title: Generation and Functional In Vitro Analysis of Semliki Forest Virus Vectors Encoding TNF-α and IFN-γ

    doi: 10.3389/fimmu.2017.01667

    Figure Lengend Snippet: Macrophages remain viable, whereas cancer cells undergo cell death after challenge with Semliki Forest virus (SFV). J774A.1 mouse macrophages and Lewis lung carcinoma (LLC) carcinoma cells were infected with SFV-DsRed particles at MOI = 10 [determined using baby hamster kidney (BHK-21) cells]. DsRed expression (first column, x -axis) was evaluated at 48 h post-infection using flow cytometry and fluorescence microscopy (scale bar 100 µM). Cell morphology was observed using bright-field microscopy (scale bar 100 µM) and cell death was quantified 48 h post-infection using flow cytometry analysis after cell staining with annexin V-FITC (second column, x -axis) and DAPI (second column, y -axis). Annexin V-positive/DAPI-negative cells were regarded as early apoptotic, whereas annexin V-positive/PI-positive cells were regarded as late apoptotic and necrotic. (A) Non-infected J774A.1 macrophages were used as a negative control. (B) J774A.1 macrophages were resistant to SFV infection, stayed viable but changed their morphology 48 h after challenge with SFV. (C) Non-infected LLC cells were used as a negative control. (D) LLC cells were susceptible to SFV infection and underwent cell death. DsRed expression was observed using microscopy in both apoptotic (white arrows) and viable (yellow arrows) cells, whereas some apoptotic cells lacked DsRed expression (red arrows). (E) The infected DsRed-positive cells were both viable and undergoing cell death. The average of duplicates with SEM

    Article Snippet: To evaluate cell viability BHK-21 and LLC cells were labeled 48 h post-infection with 1 µg/mL of fluorescein isothiocyanate (FITC)-conjugated annexin V (annexin V-FITC from Apoptosis Detection Kit; Cat. No. 556547; BD Biosciences) for 15 min followed by staining with 3 µM of DAPI (Cat. No. 422801; BioLegend, San Diego, CA, USA) for 5 min.

    Techniques: Infection, Expressing, Flow Cytometry, Cytometry, Fluorescence, Microscopy, Staining, Negative Control

    M1 macrophages activate HES1 expression and markers of differentiation in epithelial cells. HT29 cells (a) or Caco-2 cells (b) at pre-confluence were co-cultured (24h) with M1 or M2 macrophages (stained with CD18 and fluorescein isothiocyanate). In some

    Journal: Journal of Crohn's & Colitis

    Article Title: M1 Macrophages Activate Notch Signalling in Epithelial Cells: Relevance in Crohn’s Disease

    doi: 10.1093/ecco-jcc/jjw009

    Figure Lengend Snippet: M1 macrophages activate HES1 expression and markers of differentiation in epithelial cells. HT29 cells (a) or Caco-2 cells (b) at pre-confluence were co-cultured (24h) with M1 or M2 macrophages (stained with CD18 and fluorescein isothiocyanate). In some

    Article Snippet: Fluorescence-labelled (Texas red [TR] and Fluorescein isothiocyanate [FITC]) goat anti-mouse or goat anti-rabbit (1:100, Santa Cruz Biotechnology) was used as the secondary antibody, and Hoechst 33342 was added to stain the nuclei.

    Techniques: Expressing, Cell Culture, Staining

    Effects of FRAX486 and IPA3 on actin organization in WPMY-1 cells. Actin filaments were visualized by staining with FITC-coupled phalloidin, after incubation of WPMY-1 cells with DMSO, FRAX486 (1–10 μM), or IPA3 (1–10 μM) for 24 h. Shown are representative images from series with 5 independent experiments, with similar results.

    Journal: PLoS ONE

    Article Title: P21-Activated Kinase Inhibitors FRAX486 and IPA3: Inhibition of Prostate Stromal Cell Growth and Effects on Smooth Muscle Contraction in the Human Prostate

    doi: 10.1371/journal.pone.0153312

    Figure Lengend Snippet: Effects of FRAX486 and IPA3 on actin organization in WPMY-1 cells. Actin filaments were visualized by staining with FITC-coupled phalloidin, after incubation of WPMY-1 cells with DMSO, FRAX486 (1–10 μM), or IPA3 (1–10 μM) for 24 h. Shown are representative images from series with 5 independent experiments, with similar results.

    Article Snippet: Staining was performed using 100 μM FITC-labeled phalloidin (Sigma-Aldrich, Munich, Germany), according to the manufacturer’s instruction.

    Techniques: Staining, Incubation

    DDB2 reverses mesenchymal phenotype Stable pool of SCC9 cells were generated by transduction of either control retrovirus Babe-puro (SCC9:BO) or T7-DDB2 expressing retrovirus (SCC9:T7-DDB2). Expression of DDB2 was analyzed by ( A ) western blots of cell lysates (20ug) using a-tubulin as loading contrl, ( B ) DDB2 mRNA level were analyzed by qRT-PCR using cyclophilin as loading control, and ( C ) DDB2 localization was analyzed by immunocytochemical analysis with DDB2-Ab and counterstained with DAPI. ( D ) A representative phase contrast image (10×) of SCC9:B0 and SCC9:T7-DDB2 is shown. ( E ) SCC9:BO and SCC9:T7-DDB2 cells were subjected to immunocytochemical analysis using fluorophore, FITC-conjugated Phalloidin and counterstained with DAPI.

    Journal: Oncotarget

    Article Title: DDB2 regulates Epithelial-to-Mesenchymal Transition (EMT) in Oral/Head and Neck Squamous Cell Carcinoma

    doi: 10.18632/oncotarget.26168

    Figure Lengend Snippet: DDB2 reverses mesenchymal phenotype Stable pool of SCC9 cells were generated by transduction of either control retrovirus Babe-puro (SCC9:BO) or T7-DDB2 expressing retrovirus (SCC9:T7-DDB2). Expression of DDB2 was analyzed by ( A ) western blots of cell lysates (20ug) using a-tubulin as loading contrl, ( B ) DDB2 mRNA level were analyzed by qRT-PCR using cyclophilin as loading control, and ( C ) DDB2 localization was analyzed by immunocytochemical analysis with DDB2-Ab and counterstained with DAPI. ( D ) A representative phase contrast image (10×) of SCC9:B0 and SCC9:T7-DDB2 is shown. ( E ) SCC9:BO and SCC9:T7-DDB2 cells were subjected to immunocytochemical analysis using fluorophore, FITC-conjugated Phalloidin and counterstained with DAPI.

    Article Snippet: FITC conjugated Phalloidin was purchased from Sigma-Aldrich and used according to manufacturer's instructions.

    Techniques: Generated, Transduction, Expressing, Western Blot, Quantitative RT-PCR

    Confirmation of putative receptor-binding sites on the ligands using synthetic analogues by immunocytochemistry. Interaction of synthetic analogues of putative receptor-binding sites with cultured hBMECs. The interaction was detected with streptavidin-FITC conjugate. Nuclei are stained with DAPI. rDIII and rNadA (positive control) – whole recombinant ligands were incubated with hBMECs. DIII-1 – GTTYGVCSK-biotin; DIII-2 – VLIELEPPFGDSYIVVGRK-biotin; NadA-1 – AATVAIVAAYNNGQEINGFKAGETIYDIGEDGTITQK-biotin; NadA-2 – LADTDAALADTDAALDETTNALNKLGENITTFAEETK-biotin. Negative control – synthetic peptides were excluded from the assay.

    Journal: Scientific Reports

    Article Title: A simple and rapid pipeline for identification of receptor-binding sites on the surface proteins of pathogens

    doi: 10.1038/s41598-020-58305-y

    Figure Lengend Snippet: Confirmation of putative receptor-binding sites on the ligands using synthetic analogues by immunocytochemistry. Interaction of synthetic analogues of putative receptor-binding sites with cultured hBMECs. The interaction was detected with streptavidin-FITC conjugate. Nuclei are stained with DAPI. rDIII and rNadA (positive control) – whole recombinant ligands were incubated with hBMECs. DIII-1 – GTTYGVCSK-biotin; DIII-2 – VLIELEPPFGDSYIVVGRK-biotin; NadA-1 – AATVAIVAAYNNGQEINGFKAGETIYDIGEDGTITQK-biotin; NadA-2 – LADTDAALADTDAALDETTNALNKLGENITTFAEETK-biotin. Negative control – synthetic peptides were excluded from the assay.

    Article Snippet: As a positive control, rDIII and rNadA (20 µg diluted in 1 ml of PBS) were used in the assay and their binding on hBMECs was detected with anti-6x His antibody® conjugated with FITC (diluted in PBS with 1% BSA, 1:500, Abcam, UK).

    Techniques: Binding Assay, Immunocytochemistry, Cell Culture, Staining, Positive Control, Recombinant, Incubation, Negative Control

    Expression of surface molecules on NRS1 and 4-1BBL-transfected NRS1 cells. Parental NRS1 and 4-1BBL-transfected NRS1 (clone 5.13) cells were stained with either fluorescein isothiocyanate- or phycoerythrin-conjugated anti-4-1BBL, anti-CD80, anti-CD86, anti-CD54, anti-H-2K k and anti-I-A k , or an appropriate fluorochrome-conjugated immunoglobulin control. Samples were analysed by flow cytometry. Data are displayed as histograms (4-decade scale) with the immunoglobulin control as the lighter line.

    Journal: Immunology

    Article Title: Tumour rejection by gene transfer of 4-1BB ligand into a CD80+ murine squamous cell carcinoma and the requirements of co-stimulatory molecules on tumour and host cells

    doi: 10.1046/j.1365-2567.2000.t01-1-00138.x

    Figure Lengend Snippet: Expression of surface molecules on NRS1 and 4-1BBL-transfected NRS1 cells. Parental NRS1 and 4-1BBL-transfected NRS1 (clone 5.13) cells were stained with either fluorescein isothiocyanate- or phycoerythrin-conjugated anti-4-1BBL, anti-CD80, anti-CD86, anti-CD54, anti-H-2K k and anti-I-A k , or an appropriate fluorochrome-conjugated immunoglobulin control. Samples were analysed by flow cytometry. Data are displayed as histograms (4-decade scale) with the immunoglobulin control as the lighter line.

    Article Snippet: Fluorescein isothiocyanate- or phycoerythrin-conjugated anti-CD80 (16-10A1), CD86 (GL1), CD54 (3E2), H-2Kk (36-7-5) and I-Ak (11-5.2) mAbs and appropriated fluorochrome-conjugated control hamster, rat, or mouse immunoglobulin were obtained from PharMingen (San Diego, CA).

    Techniques: Expressing, Transfection, Staining, Flow Cytometry, Cytometry

    Intracellular distribution of U14. (A) Time course IFA of HST-infected Molt-3 cells harvested at the indicated time points (hpi). Acetone-fixed cells were incubated with anti-U14 and anti-p53 (DO-1) MAbs and then labeled with FITC-conjugated goat anti-mouse immunoglobulin G. (B) Confocal microscopy of HST-infected Molt-3 cells after 18 h. Anti-p53 goat PAb (FL393) and anti-U14 MAb were reacted with FITC- and tetramethyl rhodamine isothiocyanate-labeled secondary antibodies, respectively. Arrows and arrowheads indicate the cytoplasmic and nuclear dots, respectively. (C) 293 cells were transfected with the U14 expression plasmid, pEF-U14, and subjected to IFA at 24 h posttransfection. U14 was labeled with FITC-conjugated secondary antibody. Arrows and arrowheads indicate the cytoplasmic and nuclear dots, respectively. (D) Distribution of U14 and p53 in cytoplasmic and nuclear fractions. Mock (m)- and HST (i)-infected Molt-3 cells were harvested at 24 hpi and subjected to cytoplasmic and nuclear fractionation. Topoisomerase II α and α-tubulin were used as positive controls for the nuclear and cytoplasmic proteins, respectively.

    Journal: Journal of Virology

    Article Title: Human Herpesvirus 6 Open Reading Frame U14 Protein and Cellular p53 Interact with Each Other and Are Contained in the Virion

    doi: 10.1128/JVI.79.20.13037-13046.2005

    Figure Lengend Snippet: Intracellular distribution of U14. (A) Time course IFA of HST-infected Molt-3 cells harvested at the indicated time points (hpi). Acetone-fixed cells were incubated with anti-U14 and anti-p53 (DO-1) MAbs and then labeled with FITC-conjugated goat anti-mouse immunoglobulin G. (B) Confocal microscopy of HST-infected Molt-3 cells after 18 h. Anti-p53 goat PAb (FL393) and anti-U14 MAb were reacted with FITC- and tetramethyl rhodamine isothiocyanate-labeled secondary antibodies, respectively. Arrows and arrowheads indicate the cytoplasmic and nuclear dots, respectively. (C) 293 cells were transfected with the U14 expression plasmid, pEF-U14, and subjected to IFA at 24 h posttransfection. U14 was labeled with FITC-conjugated secondary antibody. Arrows and arrowheads indicate the cytoplasmic and nuclear dots, respectively. (D) Distribution of U14 and p53 in cytoplasmic and nuclear fractions. Mock (m)- and HST (i)-infected Molt-3 cells were harvested at 24 hpi and subjected to cytoplasmic and nuclear fractionation. Topoisomerase II α and α-tubulin were used as positive controls for the nuclear and cytoplasmic proteins, respectively.

    Article Snippet: The coverslips were then washed with PBS-Tween (PBST) for 5 min, followed by a wash with PBS for 5 min, and incubated for 30 min at room temperature with the appropriate secondary antibody labeled with fluorescein isothiocyanate (FITC) or tetramethyl rhodamine isothiocyanate (DAKO).

    Techniques: Immunofluorescence, Infection, Incubation, Labeling, Confocal Microscopy, Transfection, Expressing, Plasmid Preparation, Fractionation

    (a) The control cells treated with 0.1% DMSO showed no induction of apoptosis. The cells treated with OSEO showed gradual increased in percentage of apoptotic cells at concentration of (b) 50 μg/ml (16%), (c) 100 μg/ml (38%), (d) 250 μg/ml (61%), (e) 500 μg/ml (84%). (f) The resveratrol 300 μg/ml treated cells showed about 54% of apoptotic cells population. OSEO: Ocimum sanctum essential oil; MCF-7: Michigan cancer foundation-7; Annexin V-FITC: Annexin-V-fluorescein isothiocyanate; PI: Propidium iodide

    Journal: Pharmacognosy Magazine

    Article Title: Purified Essential Oil from Ocimum sanctum Linn. Triggers the Apoptotic Mechanism in Human Breast Cancer Cells

    doi: 10.4103/0973-1296.185738

    Figure Lengend Snippet: (a) The control cells treated with 0.1% DMSO showed no induction of apoptosis. The cells treated with OSEO showed gradual increased in percentage of apoptotic cells at concentration of (b) 50 μg/ml (16%), (c) 100 μg/ml (38%), (d) 250 μg/ml (61%), (e) 500 μg/ml (84%). (f) The resveratrol 300 μg/ml treated cells showed about 54% of apoptotic cells population. OSEO: Ocimum sanctum essential oil; MCF-7: Michigan cancer foundation-7; Annexin V-FITC: Annexin-V-fluorescein isothiocyanate; PI: Propidium iodide

    Article Snippet: Flow cytometry analysis (Annexin-V-fluorescein isothiocyanate) Double staining of Annexin-V-fluorescein isothiocyanate (Annexin V-FITC) and PI was performed according to the manufacturer's protocol (eBioscience, San Diego, USA).

    Techniques: Concentration Assay