1x pcr buffer  (Thermo Fisher)


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    Name:
    Universal Assay Buffer 1X
    Description:
    The ProcartaPlex Universal Assay Buffer 1X is developed for use with the ProcartaPlex assays The ProcartaPlex Universal Assay Buffer 1X is used for dilution of serum and plasma samples The ProcartaPlex Universal Assay Buffer 1X is included in our basic kits and off the shelf panels Additionally we offer the Universal Assay Buffer as a stand alone item Reported ApplicationMultiplex Immunoassay
    Catalog Number:
    epx-11111-000
    Price:
    None
    Category:
    Lab Reagents and Chemicals
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    Structured Review

    Thermo Fisher 1x pcr buffer
    The ProcartaPlex Universal Assay Buffer 1X is developed for use with the ProcartaPlex assays The ProcartaPlex Universal Assay Buffer 1X is used for dilution of serum and plasma samples The ProcartaPlex Universal Assay Buffer 1X is included in our basic kits and off the shelf panels Additionally we offer the Universal Assay Buffer as a stand alone item Reported ApplicationMultiplex Immunoassay
    https://www.bioz.com/result/1x pcr buffer/product/Thermo Fisher
    Average 99 stars, based on 667 article reviews
    Price from $9.99 to $1999.99
    1x pcr buffer - by Bioz Stars, 2020-09
    99/100 stars

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    Polymerase Chain Reaction:

    Article Title: Phylogenetics of Mycoplasma hominis clinical strains associated with gynecological infections or infertility as disclosed by an expanded multilocus sequence typing scheme
    Article Snippet: .. All amplifications were performed in a final volume of 50 µl consisting of 1X PCR buffer (10 mM Tris-HCl, pH 8.3; 50 mM KCl) (Invitrogen, USA), 2.5 mM MgCl2 (Invitrogen, USA), 0.4 µM of each primer (Sigma-Aldrich, Germany), 200 µM dNTPs (Sigma-Aldrich, Germany), 1.25 U of Taq DNA polymerase (Invitrogen, USA), and 10 µl of treated sample. .. An Applied Biosystems Thermal Cycler 2720 (Life Technologies, USA) was set up with a first cycle of denaturation for 5 min at 94 °C, followed by 35 cycles of denaturation at 94 °C for 0.45 min, annealing at 58 °C for 0.45 min, elongation at 72 °C for 1 min, and a final extension step of 7 min at 72 °C.

    Article Title: Biochemical and Molecular Analysis of Some Commercial Samples of Chilli Peppers from Mexico
    Article Snippet: .. PCR reactions were performed in a volume of 25 μ L containing 1X PCR buffer, 50 nM of MgCl2 , 200 μ M dNTP, 1.5 units of the enzyme Taq DNA polymerase (Invitrogen, Brazil), and 40–100 ng/μ L DNA samples [ ]. .. Four independent reactions were assayed using 0.4 μ M oligonucleotide OPE18 (CGGCCCACGT), or 0.2 μ M oligonucleotide MFG17 (CGCGTTCTTG), 0.2 μ M oligonucleotide MFG18 (CGGCCCACGT), or 0.2 μ M C51 (ATCAACGTACGT) and 0.2 μ M C52 (GTCGACGGACGT) oligonucleotide mixture (Invitrogen) [ , ].

    Article Title: Species identification and connectivity of marine amphipods in Canada’s three oceans
    Article Snippet: .. Polymerase chain reaction (PCR) was conducted as described in [ ]: the reaction mix contained 1X PCR buffer, 2.2 mM MgCl2 , 0.5 mM dNTPs, 0.4 μM of each primer, 1.5 U of Taq DNA polymerase (Life Technologies, Mississauga, ON, Canada), DNA template (around 40–80 ng), and water for a final volume of 25 μl. .. PCR were performed with an initial denaturation step of 3 min at 94 °C, followed by 5 cycles of 45 s at 94 °C, 45s at 46 °C, 45 s at 72 °C and 35 cycles of 45 s at 94 °C, 40 s at 51°C, 45s at 72 °C, and a final elongation step of 5 min at 72 °C.

    Protease Inhibitor:

    Article Title: Tumor-Associated APE1 Variant Exhibits Reduced Complementation Efficiency, But Does Not Promote Cancer Cell Phenotypes
    Article Snippet: .. Total cell extracts were prepared using 1X RIPA buffer with 1× Halt protease inhibitor cocktail (Pierce). .. The supernatant was transferred to a new tube, and protein concentrations were determined using either a Bradford assay (BioRad) or the BCA protein assay kit (Thermo Scientific) in combination with bovine serum albumin (BSA) to generate the standard curve.

    Incubation:

    Article Title: Contributions of Ocular Surface Components to Matrix-Metalloproteinases (MMP)-2 and MMP-9 in Feline Tears following Corneal Epithelial Wounding
    Article Snippet: .. Gels were rinsed in 1X renaturing buffer (Invitrogen) and incubated overnight in 1X developing buffer (Invitrogen) at 37°C for 18 hours. .. Gels were stained in 0.25% Coomassie Blue and destained in methanol:glacial acetic acid:distilled water (5∶1:4).

    Article Title: Somatic mutation of EZH2 (Y641) in Follicular and Diffuse Large B-cell Lymphomas of Germinal Center Origin
    Article Snippet: .. End-repair reactions were cleaned using AMPure beads and dATP was added to the 3′-ends using Klenow (exo-) 5U and 0.2mM dATP in 1X Klenow Buffer (Invitrogen) with 30-min incubation at 37°C in a Tetrad thermal cycler (MJ Research, USA). .. DNA was again cleaned on AMPure beads using a Biomek FX.

    other:

    Article Title: Dppa2/4 facilitate epigenetic remodeling during reprogramming to pluripotency
    Article Snippet: Images were taken on Leica TCS SP5 confocal and quantified using ImageJ software.

    Article Title: A Point Mutation V419L in the Sodium Channel Gene from Natural Populations of Aedes aegypti Is Involved in Resistance to λ-Cyhalothrin in Colombia
    Article Snippet: For this, we designed an allele-specific primer to 419 mutation (G1255T); and we used allele-specific primers previously developed to 1016 (G3046A) and 1534 (T4673G) mutations [ , ] ( ).

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  • 99
    Thermo Fisher phusion
    Schematics of the subcycling PCR protocol. A. Each PCR cycle involves a denaturation step and an annealing/elongation step. We introduced 4X sub-cycling the annealing/elongation step within each of the 30X amplification cycles. B. Multiplexed amplification products for pools of 7 oligos with 154–200 bp length products of varying GC content. <t>Phusion</t> and KAPA HIFI polymerases were used with and without a sub-cycling thermocycle. Each different condition is used to amplify 12 separate oligo pools with GC content ranges as follows: 1.) 16.4–34.3; 2.) 13.5–38.7; 3.) 21.5–37.3; 4.) 12.2–12.2; 5.) 12.7–40.0; 6.) 14.9–41.6; 7.) 16.4–37.6; 8.) 12.7–42.0; 9.) 20.9–40.6; 10.) 12.5–42.5; 11.) 12.7–43.7; 12.) 14.8–35.6. Results are based on electronic gels created by electrophoresis using a Perkin Elmer GX instrument with a 5k chip. *Bin shows an example of an expected PCR pattern where a strong 154-200bp product band is seen. PCR reactions were not purified and primers can be seen at the bottom of each sample.
    Phusion, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 119 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/phusion/product/Thermo Fisher
    Average 99 stars, based on 119 article reviews
    Price from $9.99 to $1999.99
    phusion - by Bioz Stars, 2020-09
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    88
    Thermo Fisher anti gpc1
    Tumor stage-specific analysis a, Table associated with receiver Operating Characteristic (ROC) curve analysis depicted in . b–f , ROC curve analysis for percent <t>GPC1</t> + crExos (red line), CA 19-9 serum levels (blue scattered line), exosomes concentration (black line) and exosomes size (scattered black line) in patients with carcinoma in situ (CIS) or stage I pancreatic cancer (n=5) (a), stage IIa pancreatic cancer (n=18) (b), stage IIb pancreatic cancer (n=117) (c), stage III pancreatic cancer (n=11) (d), and stage IV pancreas cancer (n=41) (e), compared to control (healthy donors (n=100) and patients with a benign pancreatic disease (n=26), total n=126). g . AUC: Area under the curve, CI: confidence interval. Figure 1f
    Anti Gpc1, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 88/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti gpc1/product/Thermo Fisher
    Average 88 stars, based on 2 article reviews
    Price from $9.99 to $1999.99
    anti gpc1 - by Bioz Stars, 2020-09
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    99
    Thermo Fisher pcr buffer
    <t>PCR</t> and RT-PCR of IL-17 cytokine family members in knee joint tissues at day 2 of AIA. ( a ) Representative PCR-ScreenTape (Agilent 2200 TapeStation) of IL-17 cytokine family members in tissues of WT and IL-17AKO mice. PCR analysis does not give information about expression quantity. Upper (magenta) and lower (green) markers are used as internal references to determine the molecular weight size of the sample. <t>DNA</t> ladder (25–1500 bp) on the left. GAPDH (Glyceraldehyde 3-phosphate dehydrogenase) serves as a housekeeping control gene. ( b ) Quantitative reverse transcriptase (RT)-PCR in tissues of sympathectomised IL-17AKO mice and IL-17AKO AIA control mice (n = 5 per group). mRNA level after sympathectomy are given in relation to non-sympathectomised control mice. Sympathectomy significantly (p = 0.012) decreases IL-17D-mRNA. Values are mean ± SEM. *p
    Pcr Buffer, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 5044 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/pcr buffer/product/Thermo Fisher
    Average 99 stars, based on 5044 article reviews
    Price from $9.99 to $1999.99
    pcr buffer - by Bioz Stars, 2020-09
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    Schematics of the subcycling PCR protocol. A. Each PCR cycle involves a denaturation step and an annealing/elongation step. We introduced 4X sub-cycling the annealing/elongation step within each of the 30X amplification cycles. B. Multiplexed amplification products for pools of 7 oligos with 154–200 bp length products of varying GC content. Phusion and KAPA HIFI polymerases were used with and without a sub-cycling thermocycle. Each different condition is used to amplify 12 separate oligo pools with GC content ranges as follows: 1.) 16.4–34.3; 2.) 13.5–38.7; 3.) 21.5–37.3; 4.) 12.2–12.2; 5.) 12.7–40.0; 6.) 14.9–41.6; 7.) 16.4–37.6; 8.) 12.7–42.0; 9.) 20.9–40.6; 10.) 12.5–42.5; 11.) 12.7–43.7; 12.) 14.8–35.6. Results are based on electronic gels created by electrophoresis using a Perkin Elmer GX instrument with a 5k chip. *Bin shows an example of an expected PCR pattern where a strong 154-200bp product band is seen. PCR reactions were not purified and primers can be seen at the bottom of each sample.

    Journal: PLoS ONE

    Article Title: Improved PCR Amplification of Broad Spectrum GC DNA Templates

    doi: 10.1371/journal.pone.0156478

    Figure Lengend Snippet: Schematics of the subcycling PCR protocol. A. Each PCR cycle involves a denaturation step and an annealing/elongation step. We introduced 4X sub-cycling the annealing/elongation step within each of the 30X amplification cycles. B. Multiplexed amplification products for pools of 7 oligos with 154–200 bp length products of varying GC content. Phusion and KAPA HIFI polymerases were used with and without a sub-cycling thermocycle. Each different condition is used to amplify 12 separate oligo pools with GC content ranges as follows: 1.) 16.4–34.3; 2.) 13.5–38.7; 3.) 21.5–37.3; 4.) 12.2–12.2; 5.) 12.7–40.0; 6.) 14.9–41.6; 7.) 16.4–37.6; 8.) 12.7–42.0; 9.) 20.9–40.6; 10.) 12.5–42.5; 11.) 12.7–43.7; 12.) 14.8–35.6. Results are based on electronic gels created by electrophoresis using a Perkin Elmer GX instrument with a 5k chip. *Bin shows an example of an expected PCR pattern where a strong 154-200bp product band is seen. PCR reactions were not purified and primers can be seen at the bottom of each sample.

    Article Snippet: For the Phusion PCR without additives we used 200μM dNTPs and a 1x final concentration of the Phusion 5x HF buffer from Thermo Fisher Scientific, Inc.

    Techniques: Polymerase Chain Reaction, Amplification, Electrophoresis, Chromatin Immunoprecipitation, Purification

    Tumor stage-specific analysis a, Table associated with receiver Operating Characteristic (ROC) curve analysis depicted in . b–f , ROC curve analysis for percent GPC1 + crExos (red line), CA 19-9 serum levels (blue scattered line), exosomes concentration (black line) and exosomes size (scattered black line) in patients with carcinoma in situ (CIS) or stage I pancreatic cancer (n=5) (a), stage IIa pancreatic cancer (n=18) (b), stage IIb pancreatic cancer (n=117) (c), stage III pancreatic cancer (n=11) (d), and stage IV pancreas cancer (n=41) (e), compared to control (healthy donors (n=100) and patients with a benign pancreatic disease (n=26), total n=126). g . AUC: Area under the curve, CI: confidence interval. Figure 1f

    Journal: Nature

    Article Title: Glypican1 identifies cancer exosomes and facilitates early detection of cancer

    doi: 10.1038/nature14581

    Figure Lengend Snippet: Tumor stage-specific analysis a, Table associated with receiver Operating Characteristic (ROC) curve analysis depicted in . b–f , ROC curve analysis for percent GPC1 + crExos (red line), CA 19-9 serum levels (blue scattered line), exosomes concentration (black line) and exosomes size (scattered black line) in patients with carcinoma in situ (CIS) or stage I pancreatic cancer (n=5) (a), stage IIa pancreatic cancer (n=18) (b), stage IIb pancreatic cancer (n=117) (c), stage III pancreatic cancer (n=11) (d), and stage IV pancreas cancer (n=41) (e), compared to control (healthy donors (n=100) and patients with a benign pancreatic disease (n=26), total n=126). g . AUC: Area under the curve, CI: confidence interval. Figure 1f

    Article Snippet: Exosomes-bound beads were washed 1 time in 1X PBS/2% BSA and centrifuge for 1 min at 10,000 rpm, blocked with 10% BSA with rotation at room temperature for 30 min, washed a second time in 1X PBS/2% BSA and centrifuged for 1 min at 10,000 rpm, and incubated with anti-GPC1 (PIPA528055, Thermo-Scientific, 3 μl of antibody in 20 μl of 2%BSA/1X PBS) during 30 min rotating at 4°C.

    Techniques: Concentration Assay, In Situ

    GPC1 + crExos is a non-invasive biomarker for pancreas cancer a , Percent beads with GPC1 + crExos in healthy donors (n=100), breast cancer patients (n=32) and patients with PDAC (n=190; ANOVA, post-hoc Tamhane T 2 , **** P

    Journal: Nature

    Article Title: Glypican1 identifies cancer exosomes and facilitates early detection of cancer

    doi: 10.1038/nature14581

    Figure Lengend Snippet: GPC1 + crExos is a non-invasive biomarker for pancreas cancer a , Percent beads with GPC1 + crExos in healthy donors (n=100), breast cancer patients (n=32) and patients with PDAC (n=190; ANOVA, post-hoc Tamhane T 2 , **** P

    Article Snippet: Exosomes-bound beads were washed 1 time in 1X PBS/2% BSA and centrifuge for 1 min at 10,000 rpm, blocked with 10% BSA with rotation at room temperature for 30 min, washed a second time in 1X PBS/2% BSA and centrifuged for 1 min at 10,000 rpm, and incubated with anti-GPC1 (PIPA528055, Thermo-Scientific, 3 μl of antibody in 20 μl of 2%BSA/1X PBS) during 30 min rotating at 4°C.

    Techniques: Biomarker Assay

    Exosomes isolation a , Exosomes concentration and size distribution by NanoSight® analysis of culture supernatant from NIH/3T3, MCF10A, HDF, MDA-MB-231 and E10 cells. Size mode: 105 nanometers (nm; 3 technical replicates). b , Transmission electron microscopy (TEM) micrograph of MDA-MB-231-derived exosomes. Upper right image shows a digitally zoomed inset. c , TEM micrograph of MDA-MB-231-derived exosomes following immunogold labeling for CD9. Gold particles are depicted as black dots. Upper right image shows a digitally zoomed inset. d , Immunoblot of flotillin1 and CD81 in exosomal proteins extracted from culture supernatant of E10, NIH/3T3, MDA-MB-231, MCF10A and HDF cells. e , RT–qPCR measurement of GPC1 mRNA levels in HMEL, HDF, HMLE, MCF7, MDA-MB-231, T3M4, Panc-1, MIA Paca2. Results are shown as mean ± standard deviation; n=3, 3 biological replicates, with 3 technical replicates each. f , Immunoblot of GPC1 in HMEL, HDF, HMLE, MCF7, MDA-MB-231, T3M4, Panc-1 and MIA Paca2 cell lysates (upper panel). β-actin was used as a loading control (lower panel). g , Immunoblot of GPC1 in exosomal protein lysates derived from the culture supernatant of 3 non-tumorigenic cell lines (HDF, HMEL, HMLE) and 5 tumorigenic cell lines (MCF7, MDA-MB-231, T3M4, Panc-1, MIA Paca2) (upper panel). Immunoblot of flotillin1 was used as loading control (lower panel). h , Immunoblot of flotillin1 in exosomal protein lysates from the culture supernatant of MDA-MB-231 and T3M4 following sucrose gradient purification. The protein content is assayed in each of the density layers listed.

    Journal: Nature

    Article Title: Glypican1 identifies cancer exosomes and facilitates early detection of cancer

    doi: 10.1038/nature14581

    Figure Lengend Snippet: Exosomes isolation a , Exosomes concentration and size distribution by NanoSight® analysis of culture supernatant from NIH/3T3, MCF10A, HDF, MDA-MB-231 and E10 cells. Size mode: 105 nanometers (nm; 3 technical replicates). b , Transmission electron microscopy (TEM) micrograph of MDA-MB-231-derived exosomes. Upper right image shows a digitally zoomed inset. c , TEM micrograph of MDA-MB-231-derived exosomes following immunogold labeling for CD9. Gold particles are depicted as black dots. Upper right image shows a digitally zoomed inset. d , Immunoblot of flotillin1 and CD81 in exosomal proteins extracted from culture supernatant of E10, NIH/3T3, MDA-MB-231, MCF10A and HDF cells. e , RT–qPCR measurement of GPC1 mRNA levels in HMEL, HDF, HMLE, MCF7, MDA-MB-231, T3M4, Panc-1, MIA Paca2. Results are shown as mean ± standard deviation; n=3, 3 biological replicates, with 3 technical replicates each. f , Immunoblot of GPC1 in HMEL, HDF, HMLE, MCF7, MDA-MB-231, T3M4, Panc-1 and MIA Paca2 cell lysates (upper panel). β-actin was used as a loading control (lower panel). g , Immunoblot of GPC1 in exosomal protein lysates derived from the culture supernatant of 3 non-tumorigenic cell lines (HDF, HMEL, HMLE) and 5 tumorigenic cell lines (MCF7, MDA-MB-231, T3M4, Panc-1, MIA Paca2) (upper panel). Immunoblot of flotillin1 was used as loading control (lower panel). h , Immunoblot of flotillin1 in exosomal protein lysates from the culture supernatant of MDA-MB-231 and T3M4 following sucrose gradient purification. The protein content is assayed in each of the density layers listed.

    Article Snippet: Exosomes-bound beads were washed 1 time in 1X PBS/2% BSA and centrifuge for 1 min at 10,000 rpm, blocked with 10% BSA with rotation at room temperature for 30 min, washed a second time in 1X PBS/2% BSA and centrifuged for 1 min at 10,000 rpm, and incubated with anti-GPC1 (PIPA528055, Thermo-Scientific, 3 μl of antibody in 20 μl of 2%BSA/1X PBS) during 30 min rotating at 4°C.

    Techniques: Isolation, Concentration Assay, Multiple Displacement Amplification, Transmission Assay, Electron Microscopy, Transmission Electron Microscopy, Derivative Assay, Labeling, Quantitative RT-PCR, Standard Deviation, Purification

    GPC1 + crExosomes are derived from cancer cells in tumor-bearing mice a , Longitudinal blood collection: nude mice with orthotopic MDA-MB-231 tumors (n=4 mice). b , Percentage of beads with GPC1 + exosomes harvested from systemic circulation (crExos) plotted against average tumor volume (n=4 mice, each sample analyzed in technical triplicates for GPC1). ANOVA, post-hoc Tamhane T 2 , ** P

    Journal: Nature

    Article Title: Glypican1 identifies cancer exosomes and facilitates early detection of cancer

    doi: 10.1038/nature14581

    Figure Lengend Snippet: GPC1 + crExosomes are derived from cancer cells in tumor-bearing mice a , Longitudinal blood collection: nude mice with orthotopic MDA-MB-231 tumors (n=4 mice). b , Percentage of beads with GPC1 + exosomes harvested from systemic circulation (crExos) plotted against average tumor volume (n=4 mice, each sample analyzed in technical triplicates for GPC1). ANOVA, post-hoc Tamhane T 2 , ** P

    Article Snippet: Exosomes-bound beads were washed 1 time in 1X PBS/2% BSA and centrifuge for 1 min at 10,000 rpm, blocked with 10% BSA with rotation at room temperature for 30 min, washed a second time in 1X PBS/2% BSA and centrifuged for 1 min at 10,000 rpm, and incubated with anti-GPC1 (PIPA528055, Thermo-Scientific, 3 μl of antibody in 20 μl of 2%BSA/1X PBS) during 30 min rotating at 4°C.

    Techniques: Derivative Assay, Mouse Assay, Multiple Displacement Amplification

    PDAC GEMM cross sectional studies a , Representative micrographs of H E stained pancreas from 16 days old control mice (left panel) and PKT mice presenting with (right panel, encircled) and without (middle panel) PanIN lesions. Scale bars: 100 μm. b , Ct values following qPCR analyses for oncogenic KRAS G12D , wild-type KRAS and 18S internal control RNA from exosomes of 44–48 days old PKT mice serum segregated using FACS for GPC1 + bead bound exos (red) and GPC1 − bead bound exos (blue). Data is presented as the mean ± standard deviation.

    Journal: Nature

    Article Title: Glypican1 identifies cancer exosomes and facilitates early detection of cancer

    doi: 10.1038/nature14581

    Figure Lengend Snippet: PDAC GEMM cross sectional studies a , Representative micrographs of H E stained pancreas from 16 days old control mice (left panel) and PKT mice presenting with (right panel, encircled) and without (middle panel) PanIN lesions. Scale bars: 100 μm. b , Ct values following qPCR analyses for oncogenic KRAS G12D , wild-type KRAS and 18S internal control RNA from exosomes of 44–48 days old PKT mice serum segregated using FACS for GPC1 + bead bound exos (red) and GPC1 − bead bound exos (blue). Data is presented as the mean ± standard deviation.

    Article Snippet: Exosomes-bound beads were washed 1 time in 1X PBS/2% BSA and centrifuge for 1 min at 10,000 rpm, blocked with 10% BSA with rotation at room temperature for 30 min, washed a second time in 1X PBS/2% BSA and centrifuged for 1 min at 10,000 rpm, and incubated with anti-GPC1 (PIPA528055, Thermo-Scientific, 3 μl of antibody in 20 μl of 2%BSA/1X PBS) during 30 min rotating at 4°C.

    Techniques: Staining, Mouse Assay, Real-time Polymerase Chain Reaction, FACS, Standard Deviation

    Raw scatter dot plot depicting flow cytometry analyses of beads with GPC1 + bound exosomes a , Scatter plots and histogram of flow cytometry analyses of serum exosomes on beads of a representative healthy control (left panels are secondary antibody only; right panels are GPC1 antibody and secondary antibody). b , Scatter plots and histogram of flow cytometry analysis of serum exosomes on beads of a representative pancreas cancer sample (left panels are secondary antibody only; right panels are with GPC1 antibody and secondary antibody).

    Journal: Nature

    Article Title: Glypican1 identifies cancer exosomes and facilitates early detection of cancer

    doi: 10.1038/nature14581

    Figure Lengend Snippet: Raw scatter dot plot depicting flow cytometry analyses of beads with GPC1 + bound exosomes a , Scatter plots and histogram of flow cytometry analyses of serum exosomes on beads of a representative healthy control (left panels are secondary antibody only; right panels are GPC1 antibody and secondary antibody). b , Scatter plots and histogram of flow cytometry analysis of serum exosomes on beads of a representative pancreas cancer sample (left panels are secondary antibody only; right panels are with GPC1 antibody and secondary antibody).

    Article Snippet: Exosomes-bound beads were washed 1 time in 1X PBS/2% BSA and centrifuge for 1 min at 10,000 rpm, blocked with 10% BSA with rotation at room temperature for 30 min, washed a second time in 1X PBS/2% BSA and centrifuged for 1 min at 10,000 rpm, and incubated with anti-GPC1 (PIPA528055, Thermo-Scientific, 3 μl of antibody in 20 μl of 2%BSA/1X PBS) during 30 min rotating at 4°C.

    Techniques: Flow Cytometry, Cytometry

    GPC1 + circulating exosomes predict pancreas cancer in GEMM a , Longitudinal blood collection from control and PKT mice: 4 (n=6 and n=7, respectively), 5 (n=6 and n=7), 6 (n=6 and n=6), 7 (n=6 and n=6) and 8 (n=2 and n=3) weeks of age. b , Percent beads with GPC1 + crExos from PKT (red) and control (blue) mice; ANOVA, post-hoc Tukey-Kramer test, **** P

    Journal: Nature

    Article Title: Glypican1 identifies cancer exosomes and facilitates early detection of cancer

    doi: 10.1038/nature14581

    Figure Lengend Snippet: GPC1 + circulating exosomes predict pancreas cancer in GEMM a , Longitudinal blood collection from control and PKT mice: 4 (n=6 and n=7, respectively), 5 (n=6 and n=7), 6 (n=6 and n=6), 7 (n=6 and n=6) and 8 (n=2 and n=3) weeks of age. b , Percent beads with GPC1 + crExos from PKT (red) and control (blue) mice; ANOVA, post-hoc Tukey-Kramer test, **** P

    Article Snippet: Exosomes-bound beads were washed 1 time in 1X PBS/2% BSA and centrifuge for 1 min at 10,000 rpm, blocked with 10% BSA with rotation at room temperature for 30 min, washed a second time in 1X PBS/2% BSA and centrifuged for 1 min at 10,000 rpm, and incubated with anti-GPC1 (PIPA528055, Thermo-Scientific, 3 μl of antibody in 20 μl of 2%BSA/1X PBS) during 30 min rotating at 4°C.

    Techniques: Mouse Assay

    Levels of circulating GPC1 + exosomes inform pancreas cancer resection outcome a , Longitudinal blood collection pre- (pre-op) and post-operatively (day 7). b , Percent beads with GPC1 + crExos from patients with BPD (n=4), PCPL (n=4) or PDAC (n=29) (paired two-tailed Student’s t -test, ** P

    Journal: Nature

    Article Title: Glypican1 identifies cancer exosomes and facilitates early detection of cancer

    doi: 10.1038/nature14581

    Figure Lengend Snippet: Levels of circulating GPC1 + exosomes inform pancreas cancer resection outcome a , Longitudinal blood collection pre- (pre-op) and post-operatively (day 7). b , Percent beads with GPC1 + crExos from patients with BPD (n=4), PCPL (n=4) or PDAC (n=29) (paired two-tailed Student’s t -test, ** P

    Article Snippet: Exosomes-bound beads were washed 1 time in 1X PBS/2% BSA and centrifuge for 1 min at 10,000 rpm, blocked with 10% BSA with rotation at room temperature for 30 min, washed a second time in 1X PBS/2% BSA and centrifuged for 1 min at 10,000 rpm, and incubated with anti-GPC1 (PIPA528055, Thermo-Scientific, 3 μl of antibody in 20 μl of 2%BSA/1X PBS) during 30 min rotating at 4°C.

    Techniques: Two Tailed Test

    Longitudinal human study a , Scatter plots of percent beads with GPC1 + crExos by flow cytometry in patients with pancreas cancer. Patients are divided based on metastatic disease (non-metastatic lesions, lymph node metastases and distant metastases). ANOVA, post-hoc Tukey-Kramer test, * P

    Journal: Nature

    Article Title: Glypican1 identifies cancer exosomes and facilitates early detection of cancer

    doi: 10.1038/nature14581

    Figure Lengend Snippet: Longitudinal human study a , Scatter plots of percent beads with GPC1 + crExos by flow cytometry in patients with pancreas cancer. Patients are divided based on metastatic disease (non-metastatic lesions, lymph node metastases and distant metastases). ANOVA, post-hoc Tukey-Kramer test, * P

    Article Snippet: Exosomes-bound beads were washed 1 time in 1X PBS/2% BSA and centrifuge for 1 min at 10,000 rpm, blocked with 10% BSA with rotation at room temperature for 30 min, washed a second time in 1X PBS/2% BSA and centrifuged for 1 min at 10,000 rpm, and incubated with anti-GPC1 (PIPA528055, Thermo-Scientific, 3 μl of antibody in 20 μl of 2%BSA/1X PBS) during 30 min rotating at 4°C.

    Techniques: Flow Cytometry, Cytometry

    GPC1 is present specifically on cancer exosomes a , Venn diagram of exosomal proteins from NIH/3T3 (blue), MCF 10A (red), HDF (green), E10 (yellow) and MDA-MB-231 (purple) cells. 48 proteins were exclusively detected in exosomes from MDA-MB-231 cells (n=3 protein samples, technical replicates). b , Transmission electron micrographs (TEM; left image) and immunogold TEM (right image) of GPC1. Upper right: digitally zoomed inset (n=2 experiments). c , Diagram of flow cytometry experiment to detect GPC1 on the surface of exosomes bound to beads. d , TEM of bead-bound exosomes and immunogold labeling of GPC1 (n=2 biological replicates). e , Percent beads with GPC1 + exosomes from cancer cells (red) and non-tumorigenic cells (black). f , Flow cytometry analyses of percent beads with GPC1 + exosomes from indicated cell lines (n=2 biological replicates). Negative control: secondary antibody only.

    Journal: Nature

    Article Title: Glypican1 identifies cancer exosomes and facilitates early detection of cancer

    doi: 10.1038/nature14581

    Figure Lengend Snippet: GPC1 is present specifically on cancer exosomes a , Venn diagram of exosomal proteins from NIH/3T3 (blue), MCF 10A (red), HDF (green), E10 (yellow) and MDA-MB-231 (purple) cells. 48 proteins were exclusively detected in exosomes from MDA-MB-231 cells (n=3 protein samples, technical replicates). b , Transmission electron micrographs (TEM; left image) and immunogold TEM (right image) of GPC1. Upper right: digitally zoomed inset (n=2 experiments). c , Diagram of flow cytometry experiment to detect GPC1 on the surface of exosomes bound to beads. d , TEM of bead-bound exosomes and immunogold labeling of GPC1 (n=2 biological replicates). e , Percent beads with GPC1 + exosomes from cancer cells (red) and non-tumorigenic cells (black). f , Flow cytometry analyses of percent beads with GPC1 + exosomes from indicated cell lines (n=2 biological replicates). Negative control: secondary antibody only.

    Article Snippet: Exosomes-bound beads were washed 1 time in 1X PBS/2% BSA and centrifuge for 1 min at 10,000 rpm, blocked with 10% BSA with rotation at room temperature for 30 min, washed a second time in 1X PBS/2% BSA and centrifuged for 1 min at 10,000 rpm, and incubated with anti-GPC1 (PIPA528055, Thermo-Scientific, 3 μl of antibody in 20 μl of 2%BSA/1X PBS) during 30 min rotating at 4°C.

    Techniques: Multiple Displacement Amplification, Transmission Assay, Transmission Electron Microscopy, Flow Cytometry, Cytometry, Labeling, Negative Control

    PCR and RT-PCR of IL-17 cytokine family members in knee joint tissues at day 2 of AIA. ( a ) Representative PCR-ScreenTape (Agilent 2200 TapeStation) of IL-17 cytokine family members in tissues of WT and IL-17AKO mice. PCR analysis does not give information about expression quantity. Upper (magenta) and lower (green) markers are used as internal references to determine the molecular weight size of the sample. DNA ladder (25–1500 bp) on the left. GAPDH (Glyceraldehyde 3-phosphate dehydrogenase) serves as a housekeeping control gene. ( b ) Quantitative reverse transcriptase (RT)-PCR in tissues of sympathectomised IL-17AKO mice and IL-17AKO AIA control mice (n = 5 per group). mRNA level after sympathectomy are given in relation to non-sympathectomised control mice. Sympathectomy significantly (p = 0.012) decreases IL-17D-mRNA. Values are mean ± SEM. *p

    Journal: Scientific Reports

    Article Title: Interleukin-17A is involved in mechanical hyperalgesia but not in the severity of murine antigen-induced arthritis

    doi: 10.1038/s41598-017-10509-5

    Figure Lengend Snippet: PCR and RT-PCR of IL-17 cytokine family members in knee joint tissues at day 2 of AIA. ( a ) Representative PCR-ScreenTape (Agilent 2200 TapeStation) of IL-17 cytokine family members in tissues of WT and IL-17AKO mice. PCR analysis does not give information about expression quantity. Upper (magenta) and lower (green) markers are used as internal references to determine the molecular weight size of the sample. DNA ladder (25–1500 bp) on the left. GAPDH (Glyceraldehyde 3-phosphate dehydrogenase) serves as a housekeeping control gene. ( b ) Quantitative reverse transcriptase (RT)-PCR in tissues of sympathectomised IL-17AKO mice and IL-17AKO AIA control mice (n = 5 per group). mRNA level after sympathectomy are given in relation to non-sympathectomised control mice. Sympathectomy significantly (p = 0.012) decreases IL-17D-mRNA. Values are mean ± SEM. *p

    Article Snippet: The Master Mix consisted of 1x PCR buffer, 1 unit Taq DNA polymerase per 50 µl PCR mixture, 0.2 mM dNTP mixture (10 m M dNTP Mix; Thermo Fisher Scientific), and 0.5 µM primer.

    Techniques: Polymerase Chain Reaction, Reverse Transcription Polymerase Chain Reaction, Mouse Assay, Expressing, Molecular Weight