Structured Review

Becton Dickinson facsaria cell sorter
WP1066 inhibits inducible Tregs. CD4+ cells enriched from pooled lymph nodes and spleens of C57BL/6J mice were stained with PerCP-labeled anti-CD4 (L3T4), FITC-labeled anti-CD62L (MEL-14), and APC-labeled anti-CD25 (PC61) antibodies. A <t>FACSAria</t> cell sorter
Facsaria Cell Sorter, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Images

1) Product Images from "A novel phosphorylated STAT3 inhibitor enhances T cell cytotoxicity against melanoma through inhibition of regulatory T cells"

Article Title: A novel phosphorylated STAT3 inhibitor enhances T cell cytotoxicity against melanoma through inhibition of regulatory T cells

Journal:

doi: 10.1007/s00262-008-0618-y

WP1066 inhibits inducible Tregs. CD4+ cells enriched from pooled lymph nodes and spleens of C57BL/6J mice were stained with PerCP-labeled anti-CD4 (L3T4), FITC-labeled anti-CD62L (MEL-14), and APC-labeled anti-CD25 (PC61) antibodies. A FACSAria cell sorter
Figure Legend Snippet: WP1066 inhibits inducible Tregs. CD4+ cells enriched from pooled lymph nodes and spleens of C57BL/6J mice were stained with PerCP-labeled anti-CD4 (L3T4), FITC-labeled anti-CD62L (MEL-14), and APC-labeled anti-CD25 (PC61) antibodies. A FACSAria cell sorter

Techniques Used: Mouse Assay, Staining, Labeling

Depletion of Tregs eliminates WP1066-enhanced CD3+ T cell cytotoxicity against human melanoma A375 cells. The experiment in was repeated with CD3+CD25- T cells after depleting Tregs (CD3+CD25+) with a FACSAria cell sorter. A375 cells were labeled
Figure Legend Snippet: Depletion of Tregs eliminates WP1066-enhanced CD3+ T cell cytotoxicity against human melanoma A375 cells. The experiment in was repeated with CD3+CD25- T cells after depleting Tregs (CD3+CD25+) with a FACSAria cell sorter. A375 cells were labeled

Techniques Used: Labeling

2) Product Images from "A novel phosphorylated STAT3 inhibitor enhances T cell cytotoxicity against melanoma through inhibition of regulatory T cells"

Article Title: A novel phosphorylated STAT3 inhibitor enhances T cell cytotoxicity against melanoma through inhibition of regulatory T cells

Journal:

doi: 10.1007/s00262-008-0618-y

WP1066 inhibits inducible Tregs. CD4+ cells enriched from pooled lymph nodes and spleens of C57BL/6J mice were stained with PerCP-labeled anti-CD4 (L3T4), FITC-labeled anti-CD62L (MEL-14), and APC-labeled anti-CD25 (PC61) antibodies. A FACSAria cell sorter
Figure Legend Snippet: WP1066 inhibits inducible Tregs. CD4+ cells enriched from pooled lymph nodes and spleens of C57BL/6J mice were stained with PerCP-labeled anti-CD4 (L3T4), FITC-labeled anti-CD62L (MEL-14), and APC-labeled anti-CD25 (PC61) antibodies. A FACSAria cell sorter

Techniques Used: Mouse Assay, Staining, Labeling

Depletion of Tregs eliminates WP1066-enhanced CD3+ T cell cytotoxicity against human melanoma A375 cells. The experiment in was repeated with CD3+CD25- T cells after depleting Tregs (CD3+CD25+) with a FACSAria cell sorter. A375 cells were labeled
Figure Legend Snippet: Depletion of Tregs eliminates WP1066-enhanced CD3+ T cell cytotoxicity against human melanoma A375 cells. The experiment in was repeated with CD3+CD25- T cells after depleting Tregs (CD3+CD25+) with a FACSAria cell sorter. A375 cells were labeled

Techniques Used: Labeling

Related Articles

Adoptive Transfer Assay:

Article Title: Tissue-resident memory Th17 cells maintain stable fungal commensalism in the oral mucosa
Article Snippet: A total of 106 CD4+ T cells were injected intravenously into Rag1 −/− mice, which were infected sublingually with C. albicans strain 101 on the following day. .. Adoptive transfer of C. albicans -specific Hector CD4+ T cells CD4+ T cells were first enriched from spleens of naive TCR-transgenic Hector mice with anti-CD4 microbeads (Miltenyi Biotec) following the manufacturer’s instructions and then stained with a viability marker and antibodies directed against CD3, CD4, CD90, CD44, and TCRVα2 and sorted on a FACS Aria II Cell Sorter (BD Biosciences). .. A total of 106 CD4+ TCRVα2+ CD44− Hector CD4+ T cells were injected intravenously into Rag1 −/− mice 1 day prior to sublingual infection with C. albicans strain 101.

Mouse Assay:

Article Title: Tissue-resident memory Th17 cells maintain stable fungal commensalism in the oral mucosa
Article Snippet: A total of 106 CD4+ T cells were injected intravenously into Rag1 −/− mice, which were infected sublingually with C. albicans strain 101 on the following day. .. Adoptive transfer of C. albicans -specific Hector CD4+ T cells CD4+ T cells were first enriched from spleens of naive TCR-transgenic Hector mice with anti-CD4 microbeads (Miltenyi Biotec) following the manufacturer’s instructions and then stained with a viability marker and antibodies directed against CD3, CD4, CD90, CD44, and TCRVα2 and sorted on a FACS Aria II Cell Sorter (BD Biosciences). .. A total of 106 CD4+ TCRVα2+ CD44− Hector CD4+ T cells were injected intravenously into Rag1 −/− mice 1 day prior to sublingual infection with C. albicans strain 101.

Staining:

Article Title: Tissue-resident memory Th17 cells maintain stable fungal commensalism in the oral mucosa
Article Snippet: A total of 106 CD4+ T cells were injected intravenously into Rag1 −/− mice, which were infected sublingually with C. albicans strain 101 on the following day. .. Adoptive transfer of C. albicans -specific Hector CD4+ T cells CD4+ T cells were first enriched from spleens of naive TCR-transgenic Hector mice with anti-CD4 microbeads (Miltenyi Biotec) following the manufacturer’s instructions and then stained with a viability marker and antibodies directed against CD3, CD4, CD90, CD44, and TCRVα2 and sorted on a FACS Aria II Cell Sorter (BD Biosciences). .. A total of 106 CD4+ TCRVα2+ CD44− Hector CD4+ T cells were injected intravenously into Rag1 −/− mice 1 day prior to sublingual infection with C. albicans strain 101.

Article Title: The aryl hydrocarbon receptor regulates expression of mucosal trafficking receptor GPR15
Article Snippet: The buffy coat layer was isolated and washed in HBSS containing 2% BCS. .. Cells were seeded into 24-well tissue-culture-treated plates at 1 × 106 cells per well and stimulated with plate bound anti-CD3 (clone HIT3a) and soluble anti-CD28 (clone CD28.2, both at 1 μg/mL and from Thermo Fisher Scientific) for 18–24 h. Cells were treated with AHR antagonist (CH223191) or agonist compounds pyocyanin, 1-hydro-phenazine, FICZ, TCDD, or DMSO vehicle for 24–48 h. Cells were harvested for gene expression at 24 h and flow cytometry analysis at 24–48 h. For sorting of CD4+ CD45RO+ and RO- T cells, freshly isolated human PBMC was stained with CD3-FITC, CD4-BV605, and CD45RO-APC-Cy7 (Biolegend, San Diego, CA) and acquired using FACS Aria cell sorter and FACS Diva software (BD Biosciences, San Jose, CA). ..

Marker:

Article Title: Tissue-resident memory Th17 cells maintain stable fungal commensalism in the oral mucosa
Article Snippet: A total of 106 CD4+ T cells were injected intravenously into Rag1 −/− mice, which were infected sublingually with C. albicans strain 101 on the following day. .. Adoptive transfer of C. albicans -specific Hector CD4+ T cells CD4+ T cells were first enriched from spleens of naive TCR-transgenic Hector mice with anti-CD4 microbeads (Miltenyi Biotec) following the manufacturer’s instructions and then stained with a viability marker and antibodies directed against CD3, CD4, CD90, CD44, and TCRVα2 and sorted on a FACS Aria II Cell Sorter (BD Biosciences). .. A total of 106 CD4+ TCRVα2+ CD44− Hector CD4+ T cells were injected intravenously into Rag1 −/− mice 1 day prior to sublingual infection with C. albicans strain 101.

FACS:

Article Title: Tissue-resident memory Th17 cells maintain stable fungal commensalism in the oral mucosa
Article Snippet: A total of 106 CD4+ T cells were injected intravenously into Rag1 −/− mice, which were infected sublingually with C. albicans strain 101 on the following day. .. Adoptive transfer of C. albicans -specific Hector CD4+ T cells CD4+ T cells were first enriched from spleens of naive TCR-transgenic Hector mice with anti-CD4 microbeads (Miltenyi Biotec) following the manufacturer’s instructions and then stained with a viability marker and antibodies directed against CD3, CD4, CD90, CD44, and TCRVα2 and sorted on a FACS Aria II Cell Sorter (BD Biosciences). .. A total of 106 CD4+ TCRVα2+ CD44− Hector CD4+ T cells were injected intravenously into Rag1 −/− mice 1 day prior to sublingual infection with C. albicans strain 101.

Article Title: Identification of PMN-released mutagenic factors in a co-culture model for colitis-associated cancer
Article Snippet: PMNs were activated with 0.5 nM PMA, if not stated otherwise, in cell culture medium. .. Cell sorting, co-culture, flow cytometry, analysis of mutations and calculation of mutation rates 5 × 103 –1 × 104 EGFP-negative HCT116 + chr3 or HCEC-1CT frameshift-reporter cells were sorted into 24-well plates on a FACSAria cell sorter using CloneCyt Plus software (BD Biocsiences, San Jose, CA). .. Twenty-four hours later, freshly isolated PMNs were added together with 0.5 nM PMA at effector:target ratios ranging from 0:1 (control) to 75:1.

Article Title: The aryl hydrocarbon receptor regulates expression of mucosal trafficking receptor GPR15
Article Snippet: The buffy coat layer was isolated and washed in HBSS containing 2% BCS. .. Cells were seeded into 24-well tissue-culture-treated plates at 1 × 106 cells per well and stimulated with plate bound anti-CD3 (clone HIT3a) and soluble anti-CD28 (clone CD28.2, both at 1 μg/mL and from Thermo Fisher Scientific) for 18–24 h. Cells were treated with AHR antagonist (CH223191) or agonist compounds pyocyanin, 1-hydro-phenazine, FICZ, TCDD, or DMSO vehicle for 24–48 h. Cells were harvested for gene expression at 24 h and flow cytometry analysis at 24–48 h. For sorting of CD4+ CD45RO+ and RO- T cells, freshly isolated human PBMC was stained with CD3-FITC, CD4-BV605, and CD45RO-APC-Cy7 (Biolegend, San Diego, CA) and acquired using FACS Aria cell sorter and FACS Diva software (BD Biosciences, San Jose, CA). ..

Article Title: De Novo Synthesis of Phosphatidylcholine Is Essential for the Promastigote But Not Amastigote Stage in Leishmania major
Article Snippet: For every passage, the GFP expression level was determined by flow cytometry at mid-log phase (3–7 × 106 cells/ml) using an Attune NxT Acoustic Flow Cytometer. .. After 16 passages in the presence of 50 μg/ml of GCV, cept — + pXNG4-CEPT parasites were sorted based on their GFP expression levels using a BD Biosciences FACS Aria III Plus Cell Sorter. ..

Article Title: Interleukin-33 and thymic stromal lymphopoietin, but not interleukin-25, are crucial for development of airway eosinophilia induced by chitin
Article Snippet: The cells in the negative fraction were collected and then incubated (30 min on ice) with a mixture containing PerCP/Cy5.5-conjugated anti-mouse KLRG1(Killer cell Lectin-like Receptor G1) mAb (2F1/KLRG1; BioLegend), PE/Cy7-conjugated anti-mouse CD127 mAb (A7R34; BioLegend), APC-conjugated anti-mouse Sca-1 mAb (D7; BioLegend), BD Horizon BV510-conjugated anti-mouse CD45 mAb (30-F11; BioLegend), BV421-conjugated Streptavidin (BioLegend) and eBioscience Fixable Viability Dye eFluor 780 (Thermo Fisher Scientific). .. After washing, Viability Dye-negative CD45+ lineage− Sca-1+ KLRG1+ CD127+ cells, i.e., ILC2s, were sorted using a FACS Aria II Cell Sorter (BD Biosciences) with BD FACSDiva v6.1.3 software (BD Biosciences). .. The ILC2s were resuspended in 200 μl of RPMI1640 medium supplemented with 10% FBS, 100 U/ml penicillin and 100 μg/ml streptomycin, and then cultured in the presence and absence of 50 ng/ml of each of recombinant mouse IL-25 (R & D Systems), recombinant mouse IL-33 (R & D Systems) and recombinant mouse TSLP (R & D Systems) in 96-well round-bottomed plates (680–1000 cells/well) at 37 °C in a 5.0% CO2 incubator for 2 days.

Article Title: Protein Kinase Cε, Which Is Linked to Ultraviolet Radiation-Induced Development of Squamous Cell Carcinomas, Stimulates Rapid Turnover of Adult Hair Follicle Stem Cells
Article Snippet: Keratinocytes were incubated for 1 hr in the dark at 4°C with PE-conjugated Rat Anti-Human α 6-integrin antibody at 10 μ L per 106 cells and FITC-conjugated rat antimouse CD34 antibody at 2 μ g per 106 cells (PE-α 6-integrin and FITC-CD34 antibodies; BD Biosciences). .. Keratinocyte preparations were sorted based on α 6-integrin+ and CD34 status using a FACS Aria cell sorter (BD Biosciences). .. Cells were stained with PE-conjugated anti-α6-integrin and FITC-conjugated anti-CD34 antibodies for flow cytometry.

Article Title: Interleukin-33 and thymic stromal lymphopoietin, but not interleukin-25, are crucial for development of airway eosinophilia induced by chitin
Article Snippet: For Th2 cells, the dissociated lung cells were incubated (30 min on ice) with a mixture of Ab cocktails containing APC/Cy7-conjugated anti-mouse CD45 mAb, PE-conjugated anti-mouse CD4 mAb, and APC-conjugated anti-mouse CD3ε mAb, in the presence of 7-AAD. .. After washing, the proportions of IL-5-venus+ cells among 7-AAD-negative CD45+ lineage− CD25+ ST2+ cells (as activated ILC2s) and CD45+ CD3ε+ CD4+ cells (as Th2 cells) were analyzed on a FACS Aria II Cell Sorter (BD Biosciences) with BD FACSDiva v6.1.3 software (BD Biosciences) and FlowJo v10.5.3 software (BD Biosciences; https://www.flowjo.com/solutions/flowjo). .. ILC2 culture Mice were intranasally treated with 100 μg of chitin (70–100 μm), and the lungs were collected after 24 h. Dissociated lung cells were prepared as described above and suspended in 30 ml of 30% Percoll in FACS buffer.

Co-Culture Assay:

Article Title: Identification of PMN-released mutagenic factors in a co-culture model for colitis-associated cancer
Article Snippet: PMNs were activated with 0.5 nM PMA, if not stated otherwise, in cell culture medium. .. Cell sorting, co-culture, flow cytometry, analysis of mutations and calculation of mutation rates 5 × 103 –1 × 104 EGFP-negative HCT116 + chr3 or HCEC-1CT frameshift-reporter cells were sorted into 24-well plates on a FACSAria cell sorter using CloneCyt Plus software (BD Biocsiences, San Jose, CA). .. Twenty-four hours later, freshly isolated PMNs were added together with 0.5 nM PMA at effector:target ratios ranging from 0:1 (control) to 75:1.

Flow Cytometry:

Article Title: Identification of PMN-released mutagenic factors in a co-culture model for colitis-associated cancer
Article Snippet: PMNs were activated with 0.5 nM PMA, if not stated otherwise, in cell culture medium. .. Cell sorting, co-culture, flow cytometry, analysis of mutations and calculation of mutation rates 5 × 103 –1 × 104 EGFP-negative HCT116 + chr3 or HCEC-1CT frameshift-reporter cells were sorted into 24-well plates on a FACSAria cell sorter using CloneCyt Plus software (BD Biocsiences, San Jose, CA). .. Twenty-four hours later, freshly isolated PMNs were added together with 0.5 nM PMA at effector:target ratios ranging from 0:1 (control) to 75:1.

Article Title: The aryl hydrocarbon receptor regulates expression of mucosal trafficking receptor GPR15
Article Snippet: The buffy coat layer was isolated and washed in HBSS containing 2% BCS. .. Cells were seeded into 24-well tissue-culture-treated plates at 1 × 106 cells per well and stimulated with plate bound anti-CD3 (clone HIT3a) and soluble anti-CD28 (clone CD28.2, both at 1 μg/mL and from Thermo Fisher Scientific) for 18–24 h. Cells were treated with AHR antagonist (CH223191) or agonist compounds pyocyanin, 1-hydro-phenazine, FICZ, TCDD, or DMSO vehicle for 24–48 h. Cells were harvested for gene expression at 24 h and flow cytometry analysis at 24–48 h. For sorting of CD4+ CD45RO+ and RO- T cells, freshly isolated human PBMC was stained with CD3-FITC, CD4-BV605, and CD45RO-APC-Cy7 (Biolegend, San Diego, CA) and acquired using FACS Aria cell sorter and FACS Diva software (BD Biosciences, San Jose, CA). ..

Article Title: Non-Invasive Fluorescent Monitoring of Ovarian Cancer in an Immunocompetent Mouse Model
Article Snippet: ID8 cells were grown to 70–80% confluence and then co-transfected with pROSA-puro-iRFP720 and plentiCRISPRv1 ROSA26-sgRNA2 or ROSA26-sgRNA4 using Lipofectamine 2000 (Invitrogen, CA, USA), using 3.5 µg of DNA in total. .. Transfected ID8 cells were maintained in puromycin selection for two weeks and then sorted by flow cytometry for single iRFP720+ clones ([R]730/45 vs. [R]670/30) using the FACSAria Fusion cell sorter (BD Biosciences, San Jose, CA, USA). .. Clones were maintained under selective pressure in DMEM/FBS/ITS/PS until outgrowth was evident.

Mutagenesis:

Article Title: Identification of PMN-released mutagenic factors in a co-culture model for colitis-associated cancer
Article Snippet: PMNs were activated with 0.5 nM PMA, if not stated otherwise, in cell culture medium. .. Cell sorting, co-culture, flow cytometry, analysis of mutations and calculation of mutation rates 5 × 103 –1 × 104 EGFP-negative HCT116 + chr3 or HCEC-1CT frameshift-reporter cells were sorted into 24-well plates on a FACSAria cell sorter using CloneCyt Plus software (BD Biocsiences, San Jose, CA). .. Twenty-four hours later, freshly isolated PMNs were added together with 0.5 nM PMA at effector:target ratios ranging from 0:1 (control) to 75:1.

Software:

Article Title: Identification of PMN-released mutagenic factors in a co-culture model for colitis-associated cancer
Article Snippet: PMNs were activated with 0.5 nM PMA, if not stated otherwise, in cell culture medium. .. Cell sorting, co-culture, flow cytometry, analysis of mutations and calculation of mutation rates 5 × 103 –1 × 104 EGFP-negative HCT116 + chr3 or HCEC-1CT frameshift-reporter cells were sorted into 24-well plates on a FACSAria cell sorter using CloneCyt Plus software (BD Biocsiences, San Jose, CA). .. Twenty-four hours later, freshly isolated PMNs were added together with 0.5 nM PMA at effector:target ratios ranging from 0:1 (control) to 75:1.

Article Title: The aryl hydrocarbon receptor regulates expression of mucosal trafficking receptor GPR15
Article Snippet: The buffy coat layer was isolated and washed in HBSS containing 2% BCS. .. Cells were seeded into 24-well tissue-culture-treated plates at 1 × 106 cells per well and stimulated with plate bound anti-CD3 (clone HIT3a) and soluble anti-CD28 (clone CD28.2, both at 1 μg/mL and from Thermo Fisher Scientific) for 18–24 h. Cells were treated with AHR antagonist (CH223191) or agonist compounds pyocyanin, 1-hydro-phenazine, FICZ, TCDD, or DMSO vehicle for 24–48 h. Cells were harvested for gene expression at 24 h and flow cytometry analysis at 24–48 h. For sorting of CD4+ CD45RO+ and RO- T cells, freshly isolated human PBMC was stained with CD3-FITC, CD4-BV605, and CD45RO-APC-Cy7 (Biolegend, San Diego, CA) and acquired using FACS Aria cell sorter and FACS Diva software (BD Biosciences, San Jose, CA). ..

Article Title: Interleukin-33 and thymic stromal lymphopoietin, but not interleukin-25, are crucial for development of airway eosinophilia induced by chitin
Article Snippet: The cells in the negative fraction were collected and then incubated (30 min on ice) with a mixture containing PerCP/Cy5.5-conjugated anti-mouse KLRG1(Killer cell Lectin-like Receptor G1) mAb (2F1/KLRG1; BioLegend), PE/Cy7-conjugated anti-mouse CD127 mAb (A7R34; BioLegend), APC-conjugated anti-mouse Sca-1 mAb (D7; BioLegend), BD Horizon BV510-conjugated anti-mouse CD45 mAb (30-F11; BioLegend), BV421-conjugated Streptavidin (BioLegend) and eBioscience Fixable Viability Dye eFluor 780 (Thermo Fisher Scientific). .. After washing, Viability Dye-negative CD45+ lineage− Sca-1+ KLRG1+ CD127+ cells, i.e., ILC2s, were sorted using a FACS Aria II Cell Sorter (BD Biosciences) with BD FACSDiva v6.1.3 software (BD Biosciences). .. The ILC2s were resuspended in 200 μl of RPMI1640 medium supplemented with 10% FBS, 100 U/ml penicillin and 100 μg/ml streptomycin, and then cultured in the presence and absence of 50 ng/ml of each of recombinant mouse IL-25 (R & D Systems), recombinant mouse IL-33 (R & D Systems) and recombinant mouse TSLP (R & D Systems) in 96-well round-bottomed plates (680–1000 cells/well) at 37 °C in a 5.0% CO2 incubator for 2 days.

Article Title: Interleukin-33 and thymic stromal lymphopoietin, but not interleukin-25, are crucial for development of airway eosinophilia induced by chitin
Article Snippet: For Th2 cells, the dissociated lung cells were incubated (30 min on ice) with a mixture of Ab cocktails containing APC/Cy7-conjugated anti-mouse CD45 mAb, PE-conjugated anti-mouse CD4 mAb, and APC-conjugated anti-mouse CD3ε mAb, in the presence of 7-AAD. .. After washing, the proportions of IL-5-venus+ cells among 7-AAD-negative CD45+ lineage− CD25+ ST2+ cells (as activated ILC2s) and CD45+ CD3ε+ CD4+ cells (as Th2 cells) were analyzed on a FACS Aria II Cell Sorter (BD Biosciences) with BD FACSDiva v6.1.3 software (BD Biosciences) and FlowJo v10.5.3 software (BD Biosciences; https://www.flowjo.com/solutions/flowjo). .. ILC2 culture Mice were intranasally treated with 100 μg of chitin (70–100 μm), and the lungs were collected after 24 h. Dissociated lung cells were prepared as described above and suspended in 30 ml of 30% Percoll in FACS buffer.

Expressing:

Article Title: The aryl hydrocarbon receptor regulates expression of mucosal trafficking receptor GPR15
Article Snippet: The buffy coat layer was isolated and washed in HBSS containing 2% BCS. .. Cells were seeded into 24-well tissue-culture-treated plates at 1 × 106 cells per well and stimulated with plate bound anti-CD3 (clone HIT3a) and soluble anti-CD28 (clone CD28.2, both at 1 μg/mL and from Thermo Fisher Scientific) for 18–24 h. Cells were treated with AHR antagonist (CH223191) or agonist compounds pyocyanin, 1-hydro-phenazine, FICZ, TCDD, or DMSO vehicle for 24–48 h. Cells were harvested for gene expression at 24 h and flow cytometry analysis at 24–48 h. For sorting of CD4+ CD45RO+ and RO- T cells, freshly isolated human PBMC was stained with CD3-FITC, CD4-BV605, and CD45RO-APC-Cy7 (Biolegend, San Diego, CA) and acquired using FACS Aria cell sorter and FACS Diva software (BD Biosciences, San Jose, CA). ..

Article Title: De Novo Synthesis of Phosphatidylcholine Is Essential for the Promastigote But Not Amastigote Stage in Leishmania major
Article Snippet: For every passage, the GFP expression level was determined by flow cytometry at mid-log phase (3–7 × 106 cells/ml) using an Attune NxT Acoustic Flow Cytometer. .. After 16 passages in the presence of 50 μg/ml of GCV, cept — + pXNG4-CEPT parasites were sorted based on their GFP expression levels using a BD Biosciences FACS Aria III Plus Cell Sorter. ..

Isolation:

Article Title: The aryl hydrocarbon receptor regulates expression of mucosal trafficking receptor GPR15
Article Snippet: The buffy coat layer was isolated and washed in HBSS containing 2% BCS. .. Cells were seeded into 24-well tissue-culture-treated plates at 1 × 106 cells per well and stimulated with plate bound anti-CD3 (clone HIT3a) and soluble anti-CD28 (clone CD28.2, both at 1 μg/mL and from Thermo Fisher Scientific) for 18–24 h. Cells were treated with AHR antagonist (CH223191) or agonist compounds pyocyanin, 1-hydro-phenazine, FICZ, TCDD, or DMSO vehicle for 24–48 h. Cells were harvested for gene expression at 24 h and flow cytometry analysis at 24–48 h. For sorting of CD4+ CD45RO+ and RO- T cells, freshly isolated human PBMC was stained with CD3-FITC, CD4-BV605, and CD45RO-APC-Cy7 (Biolegend, San Diego, CA) and acquired using FACS Aria cell sorter and FACS Diva software (BD Biosciences, San Jose, CA). ..

Transfection:

Article Title: Non-Invasive Fluorescent Monitoring of Ovarian Cancer in an Immunocompetent Mouse Model
Article Snippet: ID8 cells were grown to 70–80% confluence and then co-transfected with pROSA-puro-iRFP720 and plentiCRISPRv1 ROSA26-sgRNA2 or ROSA26-sgRNA4 using Lipofectamine 2000 (Invitrogen, CA, USA), using 3.5 µg of DNA in total. .. Transfected ID8 cells were maintained in puromycin selection for two weeks and then sorted by flow cytometry for single iRFP720+ clones ([R]730/45 vs. [R]670/30) using the FACSAria Fusion cell sorter (BD Biosciences, San Jose, CA, USA). .. Clones were maintained under selective pressure in DMEM/FBS/ITS/PS until outgrowth was evident.

Selection:

Article Title: Non-Invasive Fluorescent Monitoring of Ovarian Cancer in an Immunocompetent Mouse Model
Article Snippet: ID8 cells were grown to 70–80% confluence and then co-transfected with pROSA-puro-iRFP720 and plentiCRISPRv1 ROSA26-sgRNA2 or ROSA26-sgRNA4 using Lipofectamine 2000 (Invitrogen, CA, USA), using 3.5 µg of DNA in total. .. Transfected ID8 cells were maintained in puromycin selection for two weeks and then sorted by flow cytometry for single iRFP720+ clones ([R]730/45 vs. [R]670/30) using the FACSAria Fusion cell sorter (BD Biosciences, San Jose, CA, USA). .. Clones were maintained under selective pressure in DMEM/FBS/ITS/PS until outgrowth was evident.

Clone Assay:

Article Title: Non-Invasive Fluorescent Monitoring of Ovarian Cancer in an Immunocompetent Mouse Model
Article Snippet: ID8 cells were grown to 70–80% confluence and then co-transfected with pROSA-puro-iRFP720 and plentiCRISPRv1 ROSA26-sgRNA2 or ROSA26-sgRNA4 using Lipofectamine 2000 (Invitrogen, CA, USA), using 3.5 µg of DNA in total. .. Transfected ID8 cells were maintained in puromycin selection for two weeks and then sorted by flow cytometry for single iRFP720+ clones ([R]730/45 vs. [R]670/30) using the FACSAria Fusion cell sorter (BD Biosciences, San Jose, CA, USA). .. Clones were maintained under selective pressure in DMEM/FBS/ITS/PS until outgrowth was evident.

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  • 86
    Becton Dickinson facs system
    Generation and characterization of EpCAM-specific CAR-NK-92 cells. (a) Structure diagram of EpCAM-specific CAR. EF1 α , promoter; SP, signal peptide; scFv, single-chain variable fragment. (b) Transduction efficiency of lentivirus in NK-92 cells. NK-92 cells were transduced with empty lentivirus vector (Ctrl-NK-92) or lentivirus containing the EpCAM-specific CAR encoding sequence (CAR-NK-92) and sorted by a <t>FACS</t> machine with a <t>GFP</t> marker. The GFP-positive rate of transduced NK-92 cells after sorting is shown. (c) Western blot analysis of the CAR expression in NK-92 cells with a monoclonal anti-human CD3 ζ antibody. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was also detected as an internal control.
    Facs System, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/facs system/product/Becton Dickinson
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    facs system - by Bioz Stars, 2021-05
    86/100 stars
      Buy from Supplier

    86
    Becton Dickinson facsaria iii cell sorter
    Repression of activation-induced cytidine deaminase (AID) induction in germinal center B cells (GCBs) from Pten fl/fl Cγ1 Cre/+ mice. (A) Flow cytometry sorting of in vitro induced IgM-BCR expressing splenic B cells from Pten +/+ Cγ1 Cre/+ and Pten fl/fl Cγ1 Cre/+ mice. Pure splenic B cells from control and KO mice (three mice for each group) were stimulated with LPS plus interleukin-4 (IL-4) for 4 days before sorting. (B) The expression levels of PTEN, p-AKT, p-FoxO1/3a, and AID were determined in Pten +/+ Cγ1 Cre/+ and Pten fl/fl Cγ1 Cre/+ IgM-BCR expressing B cells (left). Reduced PTEN, AID expression and enhanced p-AKT and p-FoxO1/3a expression were observed in Pten fl/fl Cγ1 Cre/+ IgM-BCR expressing B cells. The in vitro induced IgM-BCR expressing B cells were isolated by the <t>FACSAria</t> <t>III</t> Cell Sorter as described in (A) . Statistical comparison of protein expression level was also shown in right. Ratio values of different proteins from Pten fl/fl Cγ1 Cre/+ IgM-BCR expressing B cells were normalized to that of the Pten +/+ Cγ1 Cre/+ IgM-BCR expressing B cells. Data were given from one representative of at least three independent experiments. * p
    Facsaria Iii Cell Sorter, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/facsaria iii cell sorter/product/Becton Dickinson
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    facsaria iii cell sorter - by Bioz Stars, 2021-05
    86/100 stars
      Buy from Supplier

    99
    Becton Dickinson facsaria fusion cell sorter
    iRFP720 fluorescence and characterisation of pROSA-iRFP720-expressing ID8 cells. ( A ) Fluorescence-activated cell sorting (FACS) enrichment of iRFP720-expressing ID8 cells. iRFP720-positive single cells ([R]730/45 vs. [R]670/30) were sorted into a 96-well plate using the <t>FACSAria</t> Fusion cell sorter (BD Biosciences, San Jose, CA, USA), and single clones were expanded and screened. ( B ) iRFP720 fluorescence was observed using the Cytation 3 imaging Multi-Mode Reader equipped with a Cy5.5 filterset (BioTek Instruments Inc., Winooski, VT, USA). Hoechst 33342 was used to visualise the nucleus (377 ex /447 em ). ( C ) Schematic diagram representing the genomic screening strategy used for identifying iRFP720 incorporation into the ROSA26 locus. ( D ) Genomic DNA isolated from pROSA-puro-iRFP720-transfected single-cell ID8 clones was PCR-amplified using a ROSA26 upstream F primer, an endogenous ROSA26 -specific R primer and an iRFP720-specific R primer, and PCR products were separated with a 1.2% agarose gel. Wild-type ROSA26 gives an 869bp product and successful iRFP720 incorporation gives a 2883bp product. C1–5: clones 1–5; WT: wild-type ID8; NTC: no-template control. ( E ) Proliferation was assessed by electrode impedance using xCELLigence real-time cell analysis. Cells (8 × 10 3 /well) were treated with cisplatin (10 µg/mL) or paclitaxel (20 nM) after 8 h, and proliferation was analysed for further 64 h. ( F ) Wild-type and pROSA-iRFP720 ID8 (3 × 10 5 cells/well) were cultured for 24 h, and then incubated for further 24 h in the presence of cisplatin (10 µg/mL) or paclitaxel (20 nM). Apoptosis was determined by Alexa Fluor®647 annexin V and PI staining on the BD LSRFortessa X-20 (BD Biosciences) flow cytometer. ( G ) Representative images of the apoptosis analysis. Data are presented as mean ± SD, n = 3.
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    Generation and characterization of EpCAM-specific CAR-NK-92 cells. (a) Structure diagram of EpCAM-specific CAR. EF1 α , promoter; SP, signal peptide; scFv, single-chain variable fragment. (b) Transduction efficiency of lentivirus in NK-92 cells. NK-92 cells were transduced with empty lentivirus vector (Ctrl-NK-92) or lentivirus containing the EpCAM-specific CAR encoding sequence (CAR-NK-92) and sorted by a FACS machine with a GFP marker. The GFP-positive rate of transduced NK-92 cells after sorting is shown. (c) Western blot analysis of the CAR expression in NK-92 cells with a monoclonal anti-human CD3 ζ antibody. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was also detected as an internal control.

    Journal: Journal of Immunology Research

    Article Title: Combination Therapy with EpCAM-CAR-NK-92 Cells and Regorafenib against Human Colorectal Cancer Models

    doi: 10.1155/2018/4263520

    Figure Lengend Snippet: Generation and characterization of EpCAM-specific CAR-NK-92 cells. (a) Structure diagram of EpCAM-specific CAR. EF1 α , promoter; SP, signal peptide; scFv, single-chain variable fragment. (b) Transduction efficiency of lentivirus in NK-92 cells. NK-92 cells were transduced with empty lentivirus vector (Ctrl-NK-92) or lentivirus containing the EpCAM-specific CAR encoding sequence (CAR-NK-92) and sorted by a FACS machine with a GFP marker. The GFP-positive rate of transduced NK-92 cells after sorting is shown. (c) Western blot analysis of the CAR expression in NK-92 cells with a monoclonal anti-human CD3 ζ antibody. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was also detected as an internal control.

    Article Snippet: After expansion for several days, GFP+ NK-92 cells were sorted with a FACS system (FACSAria III, Becton-Dickinson, USA).

    Techniques: Transduction, Plasmid Preparation, Sequencing, FACS, Marker, Western Blot, Expressing

    Repression of activation-induced cytidine deaminase (AID) induction in germinal center B cells (GCBs) from Pten fl/fl Cγ1 Cre/+ mice. (A) Flow cytometry sorting of in vitro induced IgM-BCR expressing splenic B cells from Pten +/+ Cγ1 Cre/+ and Pten fl/fl Cγ1 Cre/+ mice. Pure splenic B cells from control and KO mice (three mice for each group) were stimulated with LPS plus interleukin-4 (IL-4) for 4 days before sorting. (B) The expression levels of PTEN, p-AKT, p-FoxO1/3a, and AID were determined in Pten +/+ Cγ1 Cre/+ and Pten fl/fl Cγ1 Cre/+ IgM-BCR expressing B cells (left). Reduced PTEN, AID expression and enhanced p-AKT and p-FoxO1/3a expression were observed in Pten fl/fl Cγ1 Cre/+ IgM-BCR expressing B cells. The in vitro induced IgM-BCR expressing B cells were isolated by the FACSAria III Cell Sorter as described in (A) . Statistical comparison of protein expression level was also shown in right. Ratio values of different proteins from Pten fl/fl Cγ1 Cre/+ IgM-BCR expressing B cells were normalized to that of the Pten +/+ Cγ1 Cre/+ IgM-BCR expressing B cells. Data were given from one representative of at least three independent experiments. * p

    Journal: Frontiers in Immunology

    Article Title: PTEN-Regulated AID Transcription in Germinal Center B Cells Is Essential for the Class-Switch Recombination and IgG Antibody Responses

    doi: 10.3389/fimmu.2018.00371

    Figure Lengend Snippet: Repression of activation-induced cytidine deaminase (AID) induction in germinal center B cells (GCBs) from Pten fl/fl Cγ1 Cre/+ mice. (A) Flow cytometry sorting of in vitro induced IgM-BCR expressing splenic B cells from Pten +/+ Cγ1 Cre/+ and Pten fl/fl Cγ1 Cre/+ mice. Pure splenic B cells from control and KO mice (three mice for each group) were stimulated with LPS plus interleukin-4 (IL-4) for 4 days before sorting. (B) The expression levels of PTEN, p-AKT, p-FoxO1/3a, and AID were determined in Pten +/+ Cγ1 Cre/+ and Pten fl/fl Cγ1 Cre/+ IgM-BCR expressing B cells (left). Reduced PTEN, AID expression and enhanced p-AKT and p-FoxO1/3a expression were observed in Pten fl/fl Cγ1 Cre/+ IgM-BCR expressing B cells. The in vitro induced IgM-BCR expressing B cells were isolated by the FACSAria III Cell Sorter as described in (A) . Statistical comparison of protein expression level was also shown in right. Ratio values of different proteins from Pten fl/fl Cγ1 Cre/+ IgM-BCR expressing B cells were normalized to that of the Pten +/+ Cγ1 Cre/+ IgM-BCR expressing B cells. Data were given from one representative of at least three independent experiments. * p

    Article Snippet: Pre-incubated IgM-BCR expressing B cells were sorted by using FACSAria III Cell Sorter (BD).

    Techniques: Activation Assay, Mouse Assay, Flow Cytometry, Cytometry, In Vitro, Expressing, Isolation

    iRFP720 fluorescence and characterisation of pROSA-iRFP720-expressing ID8 cells. ( A ) Fluorescence-activated cell sorting (FACS) enrichment of iRFP720-expressing ID8 cells. iRFP720-positive single cells ([R]730/45 vs. [R]670/30) were sorted into a 96-well plate using the FACSAria Fusion cell sorter (BD Biosciences, San Jose, CA, USA), and single clones were expanded and screened. ( B ) iRFP720 fluorescence was observed using the Cytation 3 imaging Multi-Mode Reader equipped with a Cy5.5 filterset (BioTek Instruments Inc., Winooski, VT, USA). Hoechst 33342 was used to visualise the nucleus (377 ex /447 em ). ( C ) Schematic diagram representing the genomic screening strategy used for identifying iRFP720 incorporation into the ROSA26 locus. ( D ) Genomic DNA isolated from pROSA-puro-iRFP720-transfected single-cell ID8 clones was PCR-amplified using a ROSA26 upstream F primer, an endogenous ROSA26 -specific R primer and an iRFP720-specific R primer, and PCR products were separated with a 1.2% agarose gel. Wild-type ROSA26 gives an 869bp product and successful iRFP720 incorporation gives a 2883bp product. C1–5: clones 1–5; WT: wild-type ID8; NTC: no-template control. ( E ) Proliferation was assessed by electrode impedance using xCELLigence real-time cell analysis. Cells (8 × 10 3 /well) were treated with cisplatin (10 µg/mL) or paclitaxel (20 nM) after 8 h, and proliferation was analysed for further 64 h. ( F ) Wild-type and pROSA-iRFP720 ID8 (3 × 10 5 cells/well) were cultured for 24 h, and then incubated for further 24 h in the presence of cisplatin (10 µg/mL) or paclitaxel (20 nM). Apoptosis was determined by Alexa Fluor®647 annexin V and PI staining on the BD LSRFortessa X-20 (BD Biosciences) flow cytometer. ( G ) Representative images of the apoptosis analysis. Data are presented as mean ± SD, n = 3.

    Journal: Cancers

    Article Title: Non-Invasive Fluorescent Monitoring of Ovarian Cancer in an Immunocompetent Mouse Model

    doi: 10.3390/cancers11010032

    Figure Lengend Snippet: iRFP720 fluorescence and characterisation of pROSA-iRFP720-expressing ID8 cells. ( A ) Fluorescence-activated cell sorting (FACS) enrichment of iRFP720-expressing ID8 cells. iRFP720-positive single cells ([R]730/45 vs. [R]670/30) were sorted into a 96-well plate using the FACSAria Fusion cell sorter (BD Biosciences, San Jose, CA, USA), and single clones were expanded and screened. ( B ) iRFP720 fluorescence was observed using the Cytation 3 imaging Multi-Mode Reader equipped with a Cy5.5 filterset (BioTek Instruments Inc., Winooski, VT, USA). Hoechst 33342 was used to visualise the nucleus (377 ex /447 em ). ( C ) Schematic diagram representing the genomic screening strategy used for identifying iRFP720 incorporation into the ROSA26 locus. ( D ) Genomic DNA isolated from pROSA-puro-iRFP720-transfected single-cell ID8 clones was PCR-amplified using a ROSA26 upstream F primer, an endogenous ROSA26 -specific R primer and an iRFP720-specific R primer, and PCR products were separated with a 1.2% agarose gel. Wild-type ROSA26 gives an 869bp product and successful iRFP720 incorporation gives a 2883bp product. C1–5: clones 1–5; WT: wild-type ID8; NTC: no-template control. ( E ) Proliferation was assessed by electrode impedance using xCELLigence real-time cell analysis. Cells (8 × 10 3 /well) were treated with cisplatin (10 µg/mL) or paclitaxel (20 nM) after 8 h, and proliferation was analysed for further 64 h. ( F ) Wild-type and pROSA-iRFP720 ID8 (3 × 10 5 cells/well) were cultured for 24 h, and then incubated for further 24 h in the presence of cisplatin (10 µg/mL) or paclitaxel (20 nM). Apoptosis was determined by Alexa Fluor®647 annexin V and PI staining on the BD LSRFortessa X-20 (BD Biosciences) flow cytometer. ( G ) Representative images of the apoptosis analysis. Data are presented as mean ± SD, n = 3.

    Article Snippet: Transfected ID8 cells were maintained in puromycin selection for two weeks and then sorted by flow cytometry for single iRFP720+ clones ([R]730/45 vs. [R]670/30) using the FACSAria Fusion cell sorter (BD Biosciences, San Jose, CA, USA).

    Techniques: Fluorescence, Expressing, FACS, Clone Assay, Imaging, Isolation, Transfection, Polymerase Chain Reaction, Amplification, Agarose Gel Electrophoresis, Cell Culture, Incubation, Staining, Flow Cytometry