avidin biotin  (BioLegend)

 
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  • 97
    Name:
    Avidin Biotin Blocking System
    Description:
    Avidin Biotin Blocking System Apps IHC WB Size 2 x 15 ml
    Catalog Number:
    927301
    Price:
    87
    Applications:
    IHC, WB
    Conjugate:
    ANC
    Size:
    2 x 15 ml
    Category:
    Buffer Solution Chemical
    Quantity:
    1
    Buy from Supplier


    Structured Review

    BioLegend avidin biotin
    Avidin Biotin Blocking System Apps IHC WB Size 2 x 15 ml
    https://www.bioz.com/result/avidin biotin/product/BioLegend
    Average 97 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    avidin biotin - by Bioz Stars, 2020-09
    97/100 stars

    Images

    Related Articles

    Binding Assay:

    Article Title: A Structured Tumor-Immune Microenvironment in Triple Negative Breast Cancer Revealed by Multiplexed Ion Beam Imaging
    Article Snippet: .. Endogenous avidin, biotin binding proteins were blocked using Avidin/Biotin blocking systems (Biolegend, San Diego, CA). .. Sections were treated successively with avidin and biotin blocking solutions for 10 min and washed for 5 min in wash buffer.

    Article Title: MIBI-TOF: A multiplexed imaging platform relates cellular phenotypes and tissue structure
    Article Snippet: .. Endogenous avidin, biotin binding proteins were blocked using Avidin/Biotin Blocking Systems (BioLegend, San Diego, CA). .. Sections were treated successively with avidin and biotin blocking solutions for 10 min and washed for 5 min in wash buffer.

    Avidin-Biotin Assay:

    Article Title: Neonatal-derived IL-17 producing dermal γδ T cells are required to prevent spontaneous atopic dermatitis
    Article Snippet: .. Cryosections were cut to 7 um thickness, blocked in PBS + 0.3% Triton X-100 + 5% normal mouse serum for 1 hr at RT, then endogenous biotin was blocked using the Avidin/Biotin Blocking System (BioLegend) as recommended. .. Primary antibody labeling was performed in blocking buffer overnight at 4°C in a humidified chamber using the following antibodies: anti-CD4 Alexa Fluor 647 (BioLegend), goat anti-IgD purified (Cedarlane Labs), anti-GL7 Alexa Fluor 488 (BioLegend), and anti-CD11c Brilliant Violet 421 (BioLegend).

    Article Title: In vivo rescue of the hematopoietic niche by pluripotent stem cell complementation of defective osteoblast compartments
    Article Snippet: .. After washing with PBS, endogenous biotin was blocked using the Avidin/Biotin Blocking System (BioLegend). .. Sections were blocked in PBS with 10% normal donkey serum for 1 hour at RT, and then stained overnight at 4°C with chicken anti-GFP (Aves) and rat anti-CD45 biotin (BioLegend) antibodies.

    Article Title: Interleukin (IL)-18 Binding Protein Deficiency Disrupts Natural Killer Cell Maturation and Diminishes Circulating IL-18
    Article Snippet: .. Following antigen retrieval, we blocked endogenous peroxidases and phosphatases using Bloxall (Vector Labs), and blocked potential endogenous biotin with the Avidin/Biotin Blocking System (BioLegend). .. Sections were then blocked with Section Block (Electron Microscopy Sciences) to reduce non-specific binding of antibody.

    Article Title: A Structured Tumor-Immune Microenvironment in Triple Negative Breast Cancer Revealed by Multiplexed Ion Beam Imaging
    Article Snippet: .. Endogenous avidin, biotin binding proteins were blocked using Avidin/Biotin blocking systems (Biolegend, San Diego, CA). .. Sections were treated successively with avidin and biotin blocking solutions for 10 min and washed for 5 min in wash buffer.

    Article Title: MIBI-TOF: A multiplexed imaging platform relates cellular phenotypes and tissue structure
    Article Snippet: .. Endogenous avidin, biotin binding proteins were blocked using Avidin/Biotin Blocking Systems (BioLegend, San Diego, CA). .. Sections were treated successively with avidin and biotin blocking solutions for 10 min and washed for 5 min in wash buffer.

    Article Title: Neonatal-derived IL-17 producing dermal γδ T cells are required to prevent spontaneous atopic dermatitis
    Article Snippet: .. Cryosections were cut to 7 um thickness, blocked in PBS + 0.3% Triton X-100 + 5% normal mouse serum for 1 h at RT, then endogenous biotin was blocked using the Avidin/Biotin Blocking System (BioLegend) as recommended. .. Primary antibody labeling was performed in blocking buffer overnight at 4°C in a humidified chamber using the following antibodies: anti-CD4 Alexa Fluor 647 (BioLegend), goat anti-IgD purified (Cedarlane Labs), anti-GL7 Alexa Fluor 488 (BioLegend), and anti-CD11c Brilliant Violet 421 (BioLegend).

    Article Title: Multiplexed imaging of human tuberculosis granulomas uncovers immunoregulatory features conserved across tissue and blood
    Article Snippet: .. Next, all tissues underwent two rounds of blocking, the first to block endogenous biotin and avidin with an Avidin/Biotin Blocking Kit (Biolegend). .. Tissues were then washed with wash buffer and blocked for 1 hour at room temperature with 1x TBS IHC Wash Buffer with Tween 20 with 3% (v/v) normal donkey serum (Sigma-Aldrich), 0.1% (v/v) cold fish skin gelatin (Sigma Aldrich), 0.1% (v/v) Triton X-100, and 0.05% (v/v) Sodium Azide.

    Blocking Assay:

    Article Title: Neonatal-derived IL-17 producing dermal γδ T cells are required to prevent spontaneous atopic dermatitis
    Article Snippet: .. Cryosections were cut to 7 um thickness, blocked in PBS + 0.3% Triton X-100 + 5% normal mouse serum for 1 hr at RT, then endogenous biotin was blocked using the Avidin/Biotin Blocking System (BioLegend) as recommended. .. Primary antibody labeling was performed in blocking buffer overnight at 4°C in a humidified chamber using the following antibodies: anti-CD4 Alexa Fluor 647 (BioLegend), goat anti-IgD purified (Cedarlane Labs), anti-GL7 Alexa Fluor 488 (BioLegend), and anti-CD11c Brilliant Violet 421 (BioLegend).

    Article Title: In vivo rescue of the hematopoietic niche by pluripotent stem cell complementation of defective osteoblast compartments
    Article Snippet: .. After washing with PBS, endogenous biotin was blocked using the Avidin/Biotin Blocking System (BioLegend). .. Sections were blocked in PBS with 10% normal donkey serum for 1 hour at RT, and then stained overnight at 4°C with chicken anti-GFP (Aves) and rat anti-CD45 biotin (BioLegend) antibodies.

    Article Title: Interleukin (IL)-18 Binding Protein Deficiency Disrupts Natural Killer Cell Maturation and Diminishes Circulating IL-18
    Article Snippet: .. Following antigen retrieval, we blocked endogenous peroxidases and phosphatases using Bloxall (Vector Labs), and blocked potential endogenous biotin with the Avidin/Biotin Blocking System (BioLegend). .. Sections were then blocked with Section Block (Electron Microscopy Sciences) to reduce non-specific binding of antibody.

    Article Title: A Structured Tumor-Immune Microenvironment in Triple Negative Breast Cancer Revealed by Multiplexed Ion Beam Imaging
    Article Snippet: .. Endogenous avidin, biotin binding proteins were blocked using Avidin/Biotin blocking systems (Biolegend, San Diego, CA). .. Sections were treated successively with avidin and biotin blocking solutions for 10 min and washed for 5 min in wash buffer.

    Article Title: MIBI-TOF: A multiplexed imaging platform relates cellular phenotypes and tissue structure
    Article Snippet: .. Endogenous avidin, biotin binding proteins were blocked using Avidin/Biotin Blocking Systems (BioLegend, San Diego, CA). .. Sections were treated successively with avidin and biotin blocking solutions for 10 min and washed for 5 min in wash buffer.

    Article Title: Neonatal-derived IL-17 producing dermal γδ T cells are required to prevent spontaneous atopic dermatitis
    Article Snippet: .. Cryosections were cut to 7 um thickness, blocked in PBS + 0.3% Triton X-100 + 5% normal mouse serum for 1 h at RT, then endogenous biotin was blocked using the Avidin/Biotin Blocking System (BioLegend) as recommended. .. Primary antibody labeling was performed in blocking buffer overnight at 4°C in a humidified chamber using the following antibodies: anti-CD4 Alexa Fluor 647 (BioLegend), goat anti-IgD purified (Cedarlane Labs), anti-GL7 Alexa Fluor 488 (BioLegend), and anti-CD11c Brilliant Violet 421 (BioLegend).

    Article Title: Multiplexed imaging of human tuberculosis granulomas uncovers immunoregulatory features conserved across tissue and blood
    Article Snippet: .. Next, all tissues underwent two rounds of blocking, the first to block endogenous biotin and avidin with an Avidin/Biotin Blocking Kit (Biolegend). .. Tissues were then washed with wash buffer and blocked for 1 hour at room temperature with 1x TBS IHC Wash Buffer with Tween 20 with 3% (v/v) normal donkey serum (Sigma-Aldrich), 0.1% (v/v) cold fish skin gelatin (Sigma Aldrich), 0.1% (v/v) Triton X-100, and 0.05% (v/v) Sodium Azide.

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  • 85
    BioLegend biotinylated anti 4 1bb
    Expression of <t>4-1BB</t> (A) and 4-1BBL (B) in the four major thymocyte subsets during thymus regeneration. (C) The phenotypic distribution of the four thymocyte subsets during thymus regeneration. Groups of four B6 mice were killed 1, 2, 3, 4, 5, 6, 7 and 10 days after injection of cyclophosphamide. Thymocytes were stained with Pacific Blue-anti-CD4, APC-Cy7-anti-CD8, <t>biotin-anti-4-1BB,</t> biotin-anti-4-1BBL and PE-streptavidin. Data are means + SD of at least three independent experiments with four or more animals per group. * P
    Biotinylated Anti 4 1bb, supplied by BioLegend, used in various techniques. Bioz Stars score: 85/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/biotinylated anti 4 1bb/product/BioLegend
    Average 85 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    biotinylated anti 4 1bb - by Bioz Stars, 2020-09
    85/100 stars
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    90
    BioLegend biotinylated anti cd3
    The ICOS transmembrane domain mediates calcium costimulation and is required for p85 recruitment. a Surface ICOS staining of Jurkat cells stably expressing ICOS, ICOS-CD44TM, or ICOS-NT molecules. b Calcium mobilization in Jurkat cells expressing ICOS molecules of the indicated types after stimulation with <t>biotinylated</t> <t>anti-CD3</t> (0.2 μg/ml) and anti-ICOS (2 μg/ml) antibodies coligated by streptavidin. The data represent the results of three independent experiments. c Immunoblotting of phospho-ZAP70 and phospho-PLCγ in Jurkat cells expressing the indicated ICOS molecules 30 s after stimulation with anti-CD3 (0.2 μg/ml) and anti-ICOS (2 μg/ml) antibodies coligated by streptavidin. The data represent the results of three independent experiments. d Input (3% total lysates) or anti-ICOS immunoprecipitates of the indicated Jurkat cells before or 1 minute after ICOS crosslinking stimulation were probed by Western blotting for p85 and ICOS. e Immunoblotting of the indicated molecules in ICOS or ICOS-CD44TM Jurkat cells following anti-ICOS crosslinking stimulation for the indicated periods of time. Left, representative pAkt patterns; right, normalized pAkt levels of three independent experiments
    Biotinylated Anti Cd3, supplied by BioLegend, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/biotinylated anti cd3/product/BioLegend
    Average 90 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    biotinylated anti cd3 - by Bioz Stars, 2020-09
    90/100 stars
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    92
    BioLegend biotinylated anti icos antibodies
    The <t>ICOS</t> transmembrane domain mediates calcium costimulation and is required for p85 recruitment. a Surface ICOS staining of Jurkat cells stably expressing ICOS, ICOS-CD44TM, or ICOS-NT molecules. b Calcium mobilization in Jurkat cells expressing ICOS molecules of the indicated types after stimulation with <t>biotinylated</t> anti-CD3 (0.2 μg/ml) and anti-ICOS (2 μg/ml) antibodies coligated by streptavidin. The data represent the results of three independent experiments. c Immunoblotting of phospho-ZAP70 and phospho-PLCγ in Jurkat cells expressing the indicated ICOS molecules 30 s after stimulation with anti-CD3 (0.2 μg/ml) and anti-ICOS (2 μg/ml) antibodies coligated by streptavidin. The data represent the results of three independent experiments. d Input (3% total lysates) or anti-ICOS immunoprecipitates of the indicated Jurkat cells before or 1 minute after ICOS crosslinking stimulation were probed by Western blotting for p85 and ICOS. e Immunoblotting of the indicated molecules in ICOS or ICOS-CD44TM Jurkat cells following anti-ICOS crosslinking stimulation for the indicated periods of time. Left, representative pAkt patterns; right, normalized pAkt levels of three independent experiments
    Biotinylated Anti Icos Antibodies, supplied by BioLegend, used in various techniques. Bioz Stars score: 92/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/biotinylated anti icos antibodies/product/BioLegend
    Average 92 stars, based on 2 article reviews
    Price from $9.99 to $1999.99
    biotinylated anti icos antibodies - by Bioz Stars, 2020-09
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    90
    BioLegend biotinylated anti epcam antibody
    Overall workflow of the integrated nanoplatform. ( A ) First, whole-blood samples are processed for isolation of CTCs by the MagSifter. Streptavidinated 150-nm iron oxide magnetic nanoparticles (MNPs) are conjugated to <t>biotinylated</t> <t>anti-EpCAM</t> antibodies for EpCAM-positive selection of circulating tumor cells (CTCs), which are epithelial in origin. Magnetically labeled CTCs are captured on the sifter when flowed through it under an applied magnetic field and subsequently eluted in the absence of a magnetic field. ( B ) Subsequently, all eluent is directly loaded without staining onto a Nanowell device, and cells are seeded into individual compartments by centrifugation. After drying, single-cell RT-PCR mix is applied to the nanowells, which are then sealed with PCR tape. ( C ) The Nanowell device is then placed into a thermocycler for multiplex gene mutation and expression analysis via reverse transcription and PCR amplification. Multigene expression via up to four off-the-shelf hydrolysis probes with discrete emission spectra can be imaged in each nanowell. These fluorescence signals are then analyzed with custom MATLAB and R scripts for identification and characterization of putative CTCs based on modular panels of representative genes. These panels are flexible and can be adjusted to accommodate clinical needs such as therapy prediction and disease monitoring.
    Biotinylated Anti Epcam Antibody, supplied by BioLegend, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/biotinylated anti epcam antibody/product/BioLegend
    Average 90 stars, based on 1 article reviews
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    biotinylated anti epcam antibody - by Bioz Stars, 2020-09
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    Image Search Results


    Expression of 4-1BB (A) and 4-1BBL (B) in the four major thymocyte subsets during thymus regeneration. (C) The phenotypic distribution of the four thymocyte subsets during thymus regeneration. Groups of four B6 mice were killed 1, 2, 3, 4, 5, 6, 7 and 10 days after injection of cyclophosphamide. Thymocytes were stained with Pacific Blue-anti-CD4, APC-Cy7-anti-CD8, biotin-anti-4-1BB, biotin-anti-4-1BBL and PE-streptavidin. Data are means + SD of at least three independent experiments with four or more animals per group. * P

    Journal: Experimental & Molecular Medicine

    Article Title: Expression of 4-1BB and 4-1BBL in thymocytes during thymus regeneration

    doi: 10.3858/emm.2009.41.12.095

    Figure Lengend Snippet: Expression of 4-1BB (A) and 4-1BBL (B) in the four major thymocyte subsets during thymus regeneration. (C) The phenotypic distribution of the four thymocyte subsets during thymus regeneration. Groups of four B6 mice were killed 1, 2, 3, 4, 5, 6, 7 and 10 days after injection of cyclophosphamide. Thymocytes were stained with Pacific Blue-anti-CD4, APC-Cy7-anti-CD8, biotin-anti-4-1BB, biotin-anti-4-1BBL and PE-streptavidin. Data are means + SD of at least three independent experiments with four or more animals per group. * P

    Article Snippet: Biotinylated anti-4-1BB and 4-1BBL antibodies were purchased from BioLegend (San Diego, CA) and goat polyclonal anti-4-1BB and anti-4-1BBL antibodies were purchased from Santa Cruz Biotechnology (Santa Cruz, CA).

    Techniques: Expressing, Mouse Assay, Injection, Staining

    Flow cytometric analysis of the expression of 4-1BB and 4-1BBL in the mouse thymocytes. Freshly isolated thymocytes were exposed to 4-1BBL (100 ng/ml), and incubated for 6, 12, 24 and 48 h. All the cells, except the freshly isolated thymocytes, were cultured in RPMI 1640 medium for the same length of time (48 h) regardless of the duration of treatment with 4-1BBL, and stained with biotin-anti-4-1BB or biotin-anti-4-1BBL followed by PE-streptavidin. The histograms show the fluorescence intensities of 4-1BB and 4-1BBL. Data are representative of three independent experiments with similar results.

    Journal: Experimental & Molecular Medicine

    Article Title: Expression of 4-1BB and 4-1BBL in thymocytes during thymus regeneration

    doi: 10.3858/emm.2009.41.12.095

    Figure Lengend Snippet: Flow cytometric analysis of the expression of 4-1BB and 4-1BBL in the mouse thymocytes. Freshly isolated thymocytes were exposed to 4-1BBL (100 ng/ml), and incubated for 6, 12, 24 and 48 h. All the cells, except the freshly isolated thymocytes, were cultured in RPMI 1640 medium for the same length of time (48 h) regardless of the duration of treatment with 4-1BBL, and stained with biotin-anti-4-1BB or biotin-anti-4-1BBL followed by PE-streptavidin. The histograms show the fluorescence intensities of 4-1BB and 4-1BBL. Data are representative of three independent experiments with similar results.

    Article Snippet: Biotinylated anti-4-1BB and 4-1BBL antibodies were purchased from BioLegend (San Diego, CA) and goat polyclonal anti-4-1BB and anti-4-1BBL antibodies were purchased from Santa Cruz Biotechnology (Santa Cruz, CA).

    Techniques: Flow Cytometry, Expressing, Isolation, Incubation, Cell Culture, Staining, Fluorescence

    Upregulated expression of 4-1BB and 4-1BBL in the CD4 + CD8 + DP and intermediate thymocytes during thymus regeneration. B6 mice were killed in groups of four at 1, 3, 5, 7 and 10 days after injection of cyclophosphamide. They were stained with Pacific Blue-anti-CD4, APC-Cy7-anti-CD8, biotin- anti-4-1BB, biotin-anti-4-1BBL, and PE-streptavidin, and expression of 4-1BB (A) and 4-1BBL (B) was analyzed by flow cytometry in the nine thymocyte subsets. Data are the means + SD of at least three independent experiments with four or more animals in each group. * P

    Journal: Experimental & Molecular Medicine

    Article Title: Expression of 4-1BB and 4-1BBL in thymocytes during thymus regeneration

    doi: 10.3858/emm.2009.41.12.095

    Figure Lengend Snippet: Upregulated expression of 4-1BB and 4-1BBL in the CD4 + CD8 + DP and intermediate thymocytes during thymus regeneration. B6 mice were killed in groups of four at 1, 3, 5, 7 and 10 days after injection of cyclophosphamide. They were stained with Pacific Blue-anti-CD4, APC-Cy7-anti-CD8, biotin- anti-4-1BB, biotin-anti-4-1BBL, and PE-streptavidin, and expression of 4-1BB (A) and 4-1BBL (B) was analyzed by flow cytometry in the nine thymocyte subsets. Data are the means + SD of at least three independent experiments with four or more animals in each group. * P

    Article Snippet: Biotinylated anti-4-1BB and 4-1BBL antibodies were purchased from BioLegend (San Diego, CA) and goat polyclonal anti-4-1BB and anti-4-1BBL antibodies were purchased from Santa Cruz Biotechnology (Santa Cruz, CA).

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

    The ICOS transmembrane domain mediates calcium costimulation and is required for p85 recruitment. a Surface ICOS staining of Jurkat cells stably expressing ICOS, ICOS-CD44TM, or ICOS-NT molecules. b Calcium mobilization in Jurkat cells expressing ICOS molecules of the indicated types after stimulation with biotinylated anti-CD3 (0.2 μg/ml) and anti-ICOS (2 μg/ml) antibodies coligated by streptavidin. The data represent the results of three independent experiments. c Immunoblotting of phospho-ZAP70 and phospho-PLCγ in Jurkat cells expressing the indicated ICOS molecules 30 s after stimulation with anti-CD3 (0.2 μg/ml) and anti-ICOS (2 μg/ml) antibodies coligated by streptavidin. The data represent the results of three independent experiments. d Input (3% total lysates) or anti-ICOS immunoprecipitates of the indicated Jurkat cells before or 1 minute after ICOS crosslinking stimulation were probed by Western blotting for p85 and ICOS. e Immunoblotting of the indicated molecules in ICOS or ICOS-CD44TM Jurkat cells following anti-ICOS crosslinking stimulation for the indicated periods of time. Left, representative pAkt patterns; right, normalized pAkt levels of three independent experiments

    Journal: Cellular and Molecular Immunology

    Article Title: Transmembrane domain-mediated Lck association underlies bystander and costimulatory ICOS signaling

    doi: 10.1038/s41423-018-0183-z

    Figure Lengend Snippet: The ICOS transmembrane domain mediates calcium costimulation and is required for p85 recruitment. a Surface ICOS staining of Jurkat cells stably expressing ICOS, ICOS-CD44TM, or ICOS-NT molecules. b Calcium mobilization in Jurkat cells expressing ICOS molecules of the indicated types after stimulation with biotinylated anti-CD3 (0.2 μg/ml) and anti-ICOS (2 μg/ml) antibodies coligated by streptavidin. The data represent the results of three independent experiments. c Immunoblotting of phospho-ZAP70 and phospho-PLCγ in Jurkat cells expressing the indicated ICOS molecules 30 s after stimulation with anti-CD3 (0.2 μg/ml) and anti-ICOS (2 μg/ml) antibodies coligated by streptavidin. The data represent the results of three independent experiments. d Input (3% total lysates) or anti-ICOS immunoprecipitates of the indicated Jurkat cells before or 1 minute after ICOS crosslinking stimulation were probed by Western blotting for p85 and ICOS. e Immunoblotting of the indicated molecules in ICOS or ICOS-CD44TM Jurkat cells following anti-ICOS crosslinking stimulation for the indicated periods of time. Left, representative pAkt patterns; right, normalized pAkt levels of three independent experiments

    Article Snippet: Cells (2 × 106 per ml) were then incubated with biotinylated anti-CD3 (OKT3, BioLegend, 0.2 μg/ml) and biotinylated anti-ICOS antibodies (C398.4A, BioLegend, 2 μg/ml) at room temperature for 1 min. Indo-1 fluorescence was then measured for 1 min, followed by an additional 4-min recording after streptavidin was added to the cell suspension at a final concentration of 50 μg/ml.

    Techniques: Staining, Stable Transfection, Expressing, Western Blot

    The ICOS transmembrane domain, but not the cytoplasmic domain, mediates the costimulation of calcium mobilization. a Surface ICOS staining of Jurkat cells lentivirally transduced with an empty vector or vectors expressing full-length ICOS or ICOS-NT. b Example of gating GFP + stably transduced Jurkat cells and GFP − spike-in nontransduced control Jurkat cells for the calcium mobilization assay. c Calcium mobilization in Jurkat cells of the indicated types and the respective internal GFP - control cells after stimulation with biotinylated anti-CD3 (0.2 μg/ml) and anti-ICOS (2 μg/ml) antibodies coligated by streptavidin. The data represent the results of three independent experiments. d Surface ICOS staining of Jurkat cells lentivirally transduced with vectors expressing mutant ICOS molecules of the indicated types. e Surface ICOS staining after normalization of expression between ICOS-NT and ICOS-CD44TM-NT. f Calcium mobilization in Jurkat cells of the indicated types conducted as for c . The data represent the results of three independent experiments

    Journal: Cellular and Molecular Immunology

    Article Title: Transmembrane domain-mediated Lck association underlies bystander and costimulatory ICOS signaling

    doi: 10.1038/s41423-018-0183-z

    Figure Lengend Snippet: The ICOS transmembrane domain, but not the cytoplasmic domain, mediates the costimulation of calcium mobilization. a Surface ICOS staining of Jurkat cells lentivirally transduced with an empty vector or vectors expressing full-length ICOS or ICOS-NT. b Example of gating GFP + stably transduced Jurkat cells and GFP − spike-in nontransduced control Jurkat cells for the calcium mobilization assay. c Calcium mobilization in Jurkat cells of the indicated types and the respective internal GFP - control cells after stimulation with biotinylated anti-CD3 (0.2 μg/ml) and anti-ICOS (2 μg/ml) antibodies coligated by streptavidin. The data represent the results of three independent experiments. d Surface ICOS staining of Jurkat cells lentivirally transduced with vectors expressing mutant ICOS molecules of the indicated types. e Surface ICOS staining after normalization of expression between ICOS-NT and ICOS-CD44TM-NT. f Calcium mobilization in Jurkat cells of the indicated types conducted as for c . The data represent the results of three independent experiments

    Article Snippet: Cells (2 × 106 per ml) were then incubated with biotinylated anti-CD3 (OKT3, BioLegend, 0.2 μg/ml) and biotinylated anti-ICOS antibodies (C398.4A, BioLegend, 2 μg/ml) at room temperature for 1 min. Indo-1 fluorescence was then measured for 1 min, followed by an additional 4-min recording after streptavidin was added to the cell suspension at a final concentration of 50 μg/ml.

    Techniques: Staining, Transduction, Plasmid Preparation, Expressing, Stable Transfection, Calcium Mobilization Assay, Mutagenesis

    The ICOS transmembrane domain mediates calcium costimulation and is required for p85 recruitment. a Surface ICOS staining of Jurkat cells stably expressing ICOS, ICOS-CD44TM, or ICOS-NT molecules. b Calcium mobilization in Jurkat cells expressing ICOS molecules of the indicated types after stimulation with biotinylated anti-CD3 (0.2 μg/ml) and anti-ICOS (2 μg/ml) antibodies coligated by streptavidin. The data represent the results of three independent experiments. c Immunoblotting of phospho-ZAP70 and phospho-PLCγ in Jurkat cells expressing the indicated ICOS molecules 30 s after stimulation with anti-CD3 (0.2 μg/ml) and anti-ICOS (2 μg/ml) antibodies coligated by streptavidin. The data represent the results of three independent experiments. d Input (3% total lysates) or anti-ICOS immunoprecipitates of the indicated Jurkat cells before or 1 minute after ICOS crosslinking stimulation were probed by Western blotting for p85 and ICOS. e Immunoblotting of the indicated molecules in ICOS or ICOS-CD44TM Jurkat cells following anti-ICOS crosslinking stimulation for the indicated periods of time. Left, representative pAkt patterns; right, normalized pAkt levels of three independent experiments

    Journal: Cellular and Molecular Immunology

    Article Title: Transmembrane domain-mediated Lck association underlies bystander and costimulatory ICOS signaling

    doi: 10.1038/s41423-018-0183-z

    Figure Lengend Snippet: The ICOS transmembrane domain mediates calcium costimulation and is required for p85 recruitment. a Surface ICOS staining of Jurkat cells stably expressing ICOS, ICOS-CD44TM, or ICOS-NT molecules. b Calcium mobilization in Jurkat cells expressing ICOS molecules of the indicated types after stimulation with biotinylated anti-CD3 (0.2 μg/ml) and anti-ICOS (2 μg/ml) antibodies coligated by streptavidin. The data represent the results of three independent experiments. c Immunoblotting of phospho-ZAP70 and phospho-PLCγ in Jurkat cells expressing the indicated ICOS molecules 30 s after stimulation with anti-CD3 (0.2 μg/ml) and anti-ICOS (2 μg/ml) antibodies coligated by streptavidin. The data represent the results of three independent experiments. d Input (3% total lysates) or anti-ICOS immunoprecipitates of the indicated Jurkat cells before or 1 minute after ICOS crosslinking stimulation were probed by Western blotting for p85 and ICOS. e Immunoblotting of the indicated molecules in ICOS or ICOS-CD44TM Jurkat cells following anti-ICOS crosslinking stimulation for the indicated periods of time. Left, representative pAkt patterns; right, normalized pAkt levels of three independent experiments

    Article Snippet: For certain experiments, Indo-1-loaded Jurkat cells were suspended in calcium-free Ringer’s solution to incubate with biotinylated anti-CD3 and biotinylated anti-ICOS antibodies, baseline Indo-1 fluorescence was read for 1 min, streptavidin was added to a final concentration of 50 μg/ml to monitor Indo-1 fluorescence for another 2 min, and Ca2+ was added to a final concentration of 2 mM for an additional 3-min recording.

    Techniques: Staining, Stable Transfection, Expressing, Western Blot

    The ICOS transmembrane domain, but not the cytoplasmic domain, mediates the costimulation of calcium mobilization. a Surface ICOS staining of Jurkat cells lentivirally transduced with an empty vector or vectors expressing full-length ICOS or ICOS-NT. b Example of gating GFP + stably transduced Jurkat cells and GFP − spike-in nontransduced control Jurkat cells for the calcium mobilization assay. c Calcium mobilization in Jurkat cells of the indicated types and the respective internal GFP - control cells after stimulation with biotinylated anti-CD3 (0.2 μg/ml) and anti-ICOS (2 μg/ml) antibodies coligated by streptavidin. The data represent the results of three independent experiments. d Surface ICOS staining of Jurkat cells lentivirally transduced with vectors expressing mutant ICOS molecules of the indicated types. e Surface ICOS staining after normalization of expression between ICOS-NT and ICOS-CD44TM-NT. f Calcium mobilization in Jurkat cells of the indicated types conducted as for c . The data represent the results of three independent experiments

    Journal: Cellular and Molecular Immunology

    Article Title: Transmembrane domain-mediated Lck association underlies bystander and costimulatory ICOS signaling

    doi: 10.1038/s41423-018-0183-z

    Figure Lengend Snippet: The ICOS transmembrane domain, but not the cytoplasmic domain, mediates the costimulation of calcium mobilization. a Surface ICOS staining of Jurkat cells lentivirally transduced with an empty vector or vectors expressing full-length ICOS or ICOS-NT. b Example of gating GFP + stably transduced Jurkat cells and GFP − spike-in nontransduced control Jurkat cells for the calcium mobilization assay. c Calcium mobilization in Jurkat cells of the indicated types and the respective internal GFP - control cells after stimulation with biotinylated anti-CD3 (0.2 μg/ml) and anti-ICOS (2 μg/ml) antibodies coligated by streptavidin. The data represent the results of three independent experiments. d Surface ICOS staining of Jurkat cells lentivirally transduced with vectors expressing mutant ICOS molecules of the indicated types. e Surface ICOS staining after normalization of expression between ICOS-NT and ICOS-CD44TM-NT. f Calcium mobilization in Jurkat cells of the indicated types conducted as for c . The data represent the results of three independent experiments

    Article Snippet: For certain experiments, Indo-1-loaded Jurkat cells were suspended in calcium-free Ringer’s solution to incubate with biotinylated anti-CD3 and biotinylated anti-ICOS antibodies, baseline Indo-1 fluorescence was read for 1 min, streptavidin was added to a final concentration of 50 μg/ml to monitor Indo-1 fluorescence for another 2 min, and Ca2+ was added to a final concentration of 2 mM for an additional 3-min recording.

    Techniques: Staining, Transduction, Plasmid Preparation, Expressing, Stable Transfection, Calcium Mobilization Assay, Mutagenesis

    Overall workflow of the integrated nanoplatform. ( A ) First, whole-blood samples are processed for isolation of CTCs by the MagSifter. Streptavidinated 150-nm iron oxide magnetic nanoparticles (MNPs) are conjugated to biotinylated anti-EpCAM antibodies for EpCAM-positive selection of circulating tumor cells (CTCs), which are epithelial in origin. Magnetically labeled CTCs are captured on the sifter when flowed through it under an applied magnetic field and subsequently eluted in the absence of a magnetic field. ( B ) Subsequently, all eluent is directly loaded without staining onto a Nanowell device, and cells are seeded into individual compartments by centrifugation. After drying, single-cell RT-PCR mix is applied to the nanowells, which are then sealed with PCR tape. ( C ) The Nanowell device is then placed into a thermocycler for multiplex gene mutation and expression analysis via reverse transcription and PCR amplification. Multigene expression via up to four off-the-shelf hydrolysis probes with discrete emission spectra can be imaged in each nanowell. These fluorescence signals are then analyzed with custom MATLAB and R scripts for identification and characterization of putative CTCs based on modular panels of representative genes. These panels are flexible and can be adjusted to accommodate clinical needs such as therapy prediction and disease monitoring.

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

    Article Title: Molecular profiling of single circulating tumor cells from lung cancer patients

    doi: 10.1073/pnas.1608461113

    Figure Lengend Snippet: Overall workflow of the integrated nanoplatform. ( A ) First, whole-blood samples are processed for isolation of CTCs by the MagSifter. Streptavidinated 150-nm iron oxide magnetic nanoparticles (MNPs) are conjugated to biotinylated anti-EpCAM antibodies for EpCAM-positive selection of circulating tumor cells (CTCs), which are epithelial in origin. Magnetically labeled CTCs are captured on the sifter when flowed through it under an applied magnetic field and subsequently eluted in the absence of a magnetic field. ( B ) Subsequently, all eluent is directly loaded without staining onto a Nanowell device, and cells are seeded into individual compartments by centrifugation. After drying, single-cell RT-PCR mix is applied to the nanowells, which are then sealed with PCR tape. ( C ) The Nanowell device is then placed into a thermocycler for multiplex gene mutation and expression analysis via reverse transcription and PCR amplification. Multigene expression via up to four off-the-shelf hydrolysis probes with discrete emission spectra can be imaged in each nanowell. These fluorescence signals are then analyzed with custom MATLAB and R scripts for identification and characterization of putative CTCs based on modular panels of representative genes. These panels are flexible and can be adjusted to accommodate clinical needs such as therapy prediction and disease monitoring.

    Article Snippet: First, 2 mL of whole blood was mixed with 2 mL of 2× PlusCellect (R & D Systems) buffer, then incubated with 10 µL of biotinylated anti-EpCAM antibody (diluted 1:10 with PBS; BioLegend) for 1 h. Next, the sample was incubated with 20 µL of MagCellect Streptavidin Ferrofluid (MAG999; R & D Systems) for 1 h to label cells with magnetic nanoparticles.

    Techniques: Isolation, Selection, Labeling, Staining, Centrifugation, Reverse Transcription Polymerase Chain Reaction, Polymerase Chain Reaction, Multiplex Assay, Mutagenesis, Expressing, Amplification, Fluorescence