pbmc isolation  (Becton Dickinson)

 
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    Becton Dickinson pbmc isolation
    Common and personal vitamin D target genes in <t>PBMCs.</t> Based on PBMCs isolated from five individuals and treated in vitro in triplicate with 1,25(OH) 2 D 3 (Fig. 1 A, bottom) differential gene expression after RNA-seq analysis identified 568 to 1909 vitamin D target genes per individual, 327 to 598 of which are supertargets (Table S1 ). Venn diagrams were used for displaying the overlap of the respective five sets of vitamin D target genes ( A ) and supertargets ( B ). A heatmap was used, in order to compare the basal expression ( left ), FC ( center ) and 1,25(OH) 2 D 3 -induced expression ( right ) of the 91 vitamin D supertarget genes ( C ). Sorting was by their average basal expression. Genes that change their expression more than eightfold (Fig. 4 A) are highlighted in red, while those that are the drivers of common pathways (Fig. 4 B) are in green.
    Pbmc Isolation, 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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    1) Product Images from "Common and personal target genes of the micronutrient vitamin D in primary immune cells from human peripheral blood"

    Article Title: Common and personal target genes of the micronutrient vitamin D in primary immune cells from human peripheral blood

    Journal: Scientific Reports

    doi: 10.1038/s41598-020-78288-0

    Common and personal vitamin D target genes in PBMCs. Based on PBMCs isolated from five individuals and treated in vitro in triplicate with 1,25(OH) 2 D 3 (Fig. 1 A, bottom) differential gene expression after RNA-seq analysis identified 568 to 1909 vitamin D target genes per individual, 327 to 598 of which are supertargets (Table S1 ). Venn diagrams were used for displaying the overlap of the respective five sets of vitamin D target genes ( A ) and supertargets ( B ). A heatmap was used, in order to compare the basal expression ( left ), FC ( center ) and 1,25(OH) 2 D 3 -induced expression ( right ) of the 91 vitamin D supertarget genes ( C ). Sorting was by their average basal expression. Genes that change their expression more than eightfold (Fig. 4 A) are highlighted in red, while those that are the drivers of common pathways (Fig. 4 B) are in green.
    Figure Legend Snippet: Common and personal vitamin D target genes in PBMCs. Based on PBMCs isolated from five individuals and treated in vitro in triplicate with 1,25(OH) 2 D 3 (Fig. 1 A, bottom) differential gene expression after RNA-seq analysis identified 568 to 1909 vitamin D target genes per individual, 327 to 598 of which are supertargets (Table S1 ). Venn diagrams were used for displaying the overlap of the respective five sets of vitamin D target genes ( A ) and supertargets ( B ). A heatmap was used, in order to compare the basal expression ( left ), FC ( center ) and 1,25(OH) 2 D 3 -induced expression ( right ) of the 91 vitamin D supertarget genes ( C ). Sorting was by their average basal expression. Genes that change their expression more than eightfold (Fig. 4 A) are highlighted in red, while those that are the drivers of common pathways (Fig. 4 B) are in green.

    Techniques Used: Isolation, In Vitro, Expressing, RNA Sequencing Assay

    Studying gene expression in PBMCs. Design of the study ( A ). PBMCs of 12 healthy individuals (Table 1 ) were isolated and treated for 24 h with 1,25(OH) 2 D 3 (1,25D) or solvent (0.1% EtOH) in single repeat ( top ). With PBMCs of five individuals (three of which already participated in the cohort approach study) 1,25(OH) 2 D 3 and solvent treatment was performed in triplicate ( bottom ). RNA was extracted and RNA-seq analysis was performed for basal gene expression and 1,25(OH) 2 D 3 stimulated expression. A Venn diagram was used, in order to display commonly and personally expressed genes in the average of the cohort approach and in the five individuals that were investigated in triplicate ( B ).
    Figure Legend Snippet: Studying gene expression in PBMCs. Design of the study ( A ). PBMCs of 12 healthy individuals (Table 1 ) were isolated and treated for 24 h with 1,25(OH) 2 D 3 (1,25D) or solvent (0.1% EtOH) in single repeat ( top ). With PBMCs of five individuals (three of which already participated in the cohort approach study) 1,25(OH) 2 D 3 and solvent treatment was performed in triplicate ( bottom ). RNA was extracted and RNA-seq analysis was performed for basal gene expression and 1,25(OH) 2 D 3 stimulated expression. A Venn diagram was used, in order to display commonly and personally expressed genes in the average of the cohort approach and in the five individuals that were investigated in triplicate ( B ).

    Techniques Used: Expressing, Isolation, RNA Sequencing Assay

    Genome-wide view on vitamin D target genes in PBMCs. Based on PBMCs isolated from a cohort of 12 individuals and treated in vitro in a single repeat with 1,25(OH) 2 D 3 (Fig. 1 A, top) differential gene expression after RNA-seq analysis identified 877 vitamin D target genes (Table S1 ). A Venn diagram was used for displaying the overlap of this set of vitamin D target genes with those identified from a cohort of five individuals, which had been exposed in vivo to a bolus of 2000 µg vitamin D 3 16 , THP-1 cells, which had been stimulated in vitro with 1,25(OH) 2 D 3 23 , and monocytes isolated from PBMCs of a cohort of 85 individuals, which also had been stimulated in vitro with 1,25(OH) 2 D 3 39 ( A ). A Manhattan plot displays the genome-wide distribution of the 877 vitamin D target genes and indicates their responsiveness ( B ). The 10 most responsive genes (FC > 16) are highlighted.
    Figure Legend Snippet: Genome-wide view on vitamin D target genes in PBMCs. Based on PBMCs isolated from a cohort of 12 individuals and treated in vitro in a single repeat with 1,25(OH) 2 D 3 (Fig. 1 A, top) differential gene expression after RNA-seq analysis identified 877 vitamin D target genes (Table S1 ). A Venn diagram was used for displaying the overlap of this set of vitamin D target genes with those identified from a cohort of five individuals, which had been exposed in vivo to a bolus of 2000 µg vitamin D 3 16 , THP-1 cells, which had been stimulated in vitro with 1,25(OH) 2 D 3 23 , and monocytes isolated from PBMCs of a cohort of 85 individuals, which also had been stimulated in vitro with 1,25(OH) 2 D 3 39 ( A ). A Manhattan plot displays the genome-wide distribution of the 877 vitamin D target genes and indicates their responsiveness ( B ). The 10 most responsive genes (FC > 16) are highlighted.

    Techniques Used: Genome Wide, Isolation, In Vitro, Expressing, RNA Sequencing Assay, In Vivo

    2) Product Images from "Thiazolides Elicit Anti-Viral Innate Immunity and Reduce HIV Replication"

    Article Title: Thiazolides Elicit Anti-Viral Innate Immunity and Reduce HIV Replication

    Journal: Scientific Reports

    doi: 10.1038/srep27148

    ( a ) p24 viral antigen levels 10 days post-infection in the supernatant of HIV-1 infected PBMCs treated with TIZ (10 μg/mL) or RM-4848 (10 μg/mL). Results obtained with PBMC of 20 different healthy donors are shown. Median values and statistical significance is presented. ( b ) Inhibition (%) of viral replication in PBMC of 20 different healthy donors infected in vitro with HIV-1 in the presence of TIZ- (10 μg/mL) or RM-4848- (10 μg/mL). Median values and statistical significance is presented. ( c ) p24 viral antigen levels in PBMCs infected with HIV-1 in the presence of RM-4848 throughout infection (RM-4848)(10 μg/mL) or in medium alone (no treatment). In addition to these two situations, results of 3 other experimental conditions are shown: 1) PBMCs treated with RM-4848 before being infected (pre-infection); 2) RM-4848 present during the 24 hours infection period (during infection) and then washed away; 3) RM-4848 added 24 hours after infection (post infection). Mean values ± standard errors are shown.
    Figure Legend Snippet: ( a ) p24 viral antigen levels 10 days post-infection in the supernatant of HIV-1 infected PBMCs treated with TIZ (10 μg/mL) or RM-4848 (10 μg/mL). Results obtained with PBMC of 20 different healthy donors are shown. Median values and statistical significance is presented. ( b ) Inhibition (%) of viral replication in PBMC of 20 different healthy donors infected in vitro with HIV-1 in the presence of TIZ- (10 μg/mL) or RM-4848- (10 μg/mL). Median values and statistical significance is presented. ( c ) p24 viral antigen levels in PBMCs infected with HIV-1 in the presence of RM-4848 throughout infection (RM-4848)(10 μg/mL) or in medium alone (no treatment). In addition to these two situations, results of 3 other experimental conditions are shown: 1) PBMCs treated with RM-4848 before being infected (pre-infection); 2) RM-4848 present during the 24 hours infection period (during infection) and then washed away; 3) RM-4848 added 24 hours after infection (post infection). Mean values ± standard errors are shown.

    Techniques Used: Infection, Inhibition, In Vitro

    Cytokines (IL-2, IFNγ) and chemokines (MCP-1, MIP-1α, MIP-1β, RANTES) production (pg/ml) by PBMCs of healthy donors. PBMCs were either uninfected (white bars) or HIV-1-infected (grey bars) and were cultured in medium alone (no drug), or in medium containing either TIZ- (10 μg/mL) ( a ) or RM-4848- (10 μg/mL) ( b ). Mean values 7 days post-infection ± standard errors and statistical differences are shown.
    Figure Legend Snippet: Cytokines (IL-2, IFNγ) and chemokines (MCP-1, MIP-1α, MIP-1β, RANTES) production (pg/ml) by PBMCs of healthy donors. PBMCs were either uninfected (white bars) or HIV-1-infected (grey bars) and were cultured in medium alone (no drug), or in medium containing either TIZ- (10 μg/mL) ( a ) or RM-4848- (10 μg/mL) ( b ). Mean values 7 days post-infection ± standard errors and statistical differences are shown.

    Techniques Used: Infection, Cell Culture

    mRNA expression of 84 genes that are part of the HIV-1 life cycle and known to directly obstacle HIV infectivity has been assessed by real-time quantitative RT-PCR, calculated relative to five housekeeping genes and shown as fold-change expression from the untreated sample. Gene expression (nfold) is shown as a color scale from green to red (−2 to +9) (MEV multiple experiment viewer software). Results were obtained in HIV1-infected PBMCs at day 7 post infection, cultured in medium containing TIZ (10 μg/mL) or RM-4848 (10 μg/mL). Only targets showing > 2-fold modulation are considered significant and are shown in tab.
    Figure Legend Snippet: mRNA expression of 84 genes that are part of the HIV-1 life cycle and known to directly obstacle HIV infectivity has been assessed by real-time quantitative RT-PCR, calculated relative to five housekeeping genes and shown as fold-change expression from the untreated sample. Gene expression (nfold) is shown as a color scale from green to red (−2 to +9) (MEV multiple experiment viewer software). Results were obtained in HIV1-infected PBMCs at day 7 post infection, cultured in medium containing TIZ (10 μg/mL) or RM-4848 (10 μg/mL). Only targets showing > 2-fold modulation are considered significant and are shown in tab.

    Techniques Used: Expressing, Infection, Quantitative RT-PCR, Software, Cell Culture

    mRNA expression of genes involved in cholesterol metabolism and efflux in PBMCs of healthy donors. ( a ) Uninfected PBMCs cultured in medium containing TIZ (10 μg/mL); ( b ) Uninfected PBMCs cultured in medium containing RM-4848 (10 μg/mL); ( c ) HIV-1–infected PBMCs (day 7 post infection) cultured in medium containing TIZ (10 μg/mL); ( d ) HIV-1–infected PBMCs (day 7 post infection) cultured in medium containing RM-4848 (10 μg/mL). Gene expression genes is assessed by quantitative RT-PCR and shown as fold-change expression from the untreated sample. Only targets showing > 2-fold modulation are considered significant. Mean values ± S.D. and p values are indicated.
    Figure Legend Snippet: mRNA expression of genes involved in cholesterol metabolism and efflux in PBMCs of healthy donors. ( a ) Uninfected PBMCs cultured in medium containing TIZ (10 μg/mL); ( b ) Uninfected PBMCs cultured in medium containing RM-4848 (10 μg/mL); ( c ) HIV-1–infected PBMCs (day 7 post infection) cultured in medium containing TIZ (10 μg/mL); ( d ) HIV-1–infected PBMCs (day 7 post infection) cultured in medium containing RM-4848 (10 μg/mL). Gene expression genes is assessed by quantitative RT-PCR and shown as fold-change expression from the untreated sample. Only targets showing > 2-fold modulation are considered significant. Mean values ± S.D. and p values are indicated.

    Techniques Used: Expressing, Cell Culture, Infection, Quantitative RT-PCR

    p24 viral antigen concentration in HIV-1 - infected PBMCs cultured in medium alone (no drug) or in the presence of TO-901317 (1 μM), an LXR agonist. Results were obtained with PBMCs of 20 different healthy donors 10 days post-infection. Mean values ± standard errors and statistical significance are shown.
    Figure Legend Snippet: p24 viral antigen concentration in HIV-1 - infected PBMCs cultured in medium alone (no drug) or in the presence of TO-901317 (1 μM), an LXR agonist. Results were obtained with PBMCs of 20 different healthy donors 10 days post-infection. Mean values ± standard errors and statistical significance are shown.

    Techniques Used: Concentration Assay, Infection, Cell Culture

    3) Product Images from "A Regulatory Polymorphism in HAVCR2 Modulates Susceptibility to HIV-1 Infection"

    Article Title: A Regulatory Polymorphism in HAVCR2 Modulates Susceptibility to HIV-1 Infection

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0106442

    Box-and-whisker plot of HAVCR2 expression depending on rs4704846 genotype. Data derive from PBMCs from 40 healthy volunteers uninfected or infected with HIV-1. HAVCR2 transcript levels are log-transformed and shown in standard box-and-whisker plot representation (thick line: median; box: quartiles; whiskers: 1.5 × interquartile range); p values are calculated using the Student's t -test.
    Figure Legend Snippet: Box-and-whisker plot of HAVCR2 expression depending on rs4704846 genotype. Data derive from PBMCs from 40 healthy volunteers uninfected or infected with HIV-1. HAVCR2 transcript levels are log-transformed and shown in standard box-and-whisker plot representation (thick line: median; box: quartiles; whiskers: 1.5 × interquartile range); p values are calculated using the Student's t -test.

    Techniques Used: Whisker Assay, Expressing, Infection, Transformation Assay

    4) Product Images from "Focal Irradiation And Systemic Transforming Growth Factor β Blockade in Metastatic Breast Cancer"

    Article Title: Focal Irradiation And Systemic Transforming Growth Factor β Blockade in Metastatic Breast Cancer

    Journal: Clinical cancer research : an official journal of the American Association for Cancer Research

    doi: 10.1158/1078-0432.CCR-17-3322

    Effect of TGFβ blockade on PBMC levels, survivin-reactive CD8 + T cells, and memory T cells (A) Individual log2 fold changes in PBMC levels relative to baseline in patients receiving 1 or 10mg/kg fresolimumab. (B) Tetramer binding data are shown as % survivin-positive CD8+ T cells over the course of a 15 week treatment. The presumed threshold of median + IQR of n=11 healthy control levels is indicated in gray. (N=NYU patient; U=UCLA patient; black=1mg and red=10mg fresolimumab, green=11 healthy donors. N01 and N01* indicates repeated draws at week 0 and week 2 due to significant treatment delay). (C-F) T cell differentiation was assessed within each CD4+ or CD8+ T cell pool giving naïve (N, CCR7+CD45RA+), central memory (CM, CCR7+CD45RA-), effector memory (EF, CCR7-CD45RA-) and effector cells (E, CCR7-CD45RA+) T cells. Data are shown as individual baseline values ranked according to survival in C) the CD8+ compartment or D) the CD4+ compartment or as log2 fold change to baseline in E and F respectively.
    Figure Legend Snippet: Effect of TGFβ blockade on PBMC levels, survivin-reactive CD8 + T cells, and memory T cells (A) Individual log2 fold changes in PBMC levels relative to baseline in patients receiving 1 or 10mg/kg fresolimumab. (B) Tetramer binding data are shown as % survivin-positive CD8+ T cells over the course of a 15 week treatment. The presumed threshold of median + IQR of n=11 healthy control levels is indicated in gray. (N=NYU patient; U=UCLA patient; black=1mg and red=10mg fresolimumab, green=11 healthy donors. N01 and N01* indicates repeated draws at week 0 and week 2 due to significant treatment delay). (C-F) T cell differentiation was assessed within each CD4+ or CD8+ T cell pool giving naïve (N, CCR7+CD45RA+), central memory (CM, CCR7+CD45RA-), effector memory (EF, CCR7-CD45RA-) and effector cells (E, CCR7-CD45RA+) T cells. Data are shown as individual baseline values ranked according to survival in C) the CD8+ compartment or D) the CD4+ compartment or as log2 fold change to baseline in E and F respectively.

    Techniques Used: Binding Assay, Cell Differentiation

    Related Articles

    Isolation:

    Article Title: Biologically driven cut-off definition of lymphocyte ratios in metastatic breast cancer and association with exosomal subpopulations and prognosis
    Article Snippet: Exosome internalization experiments To compare the ability of CD14 + monocytes and CD3 + lymphocytes to internalize plasma-derived exosomes, 1.5 × 1010 exosomes plasma-derived were resuspended in PBS and stained with 5 μM DiD for 30 minutes at 37 °C. .. 2 × 105 isolated PBMCs were then incubated with DiD-labelled vesicles for 5 and 24 hours, respectively, and subsequently stained for CD45, CD3 and CD14 (Becton Dickinson) and analyzed by flow cytometry (FACSCanto II, Becton Dickinson) by gating either on CD14 + or CD3 + PBMCs. ..

    Article Title: Anti-inflammatory effects of α-MSH through p-CREB expression in sarcoidosis like granuloma model
    Article Snippet: .. PBMC isolation PBMC were isolated from whole blood using a fully-closed system with Ficoll™ Hypaque™ Solution offered by BD Biosciences, USA (Vacutainer® CPT™ Mononuclear Cell Preparation Tube -Sodium Citrate) per manufacturer recommendation and previous studies protocols , . ..

    Article Title: Expression of Toll-Like Receptors 3, 7, and 9 in Peripheral Blood Mononuclear Cells from Patients with Systemic Lupus Erythematosus
    Article Snippet: .. Assessment of TLR Expression in PBMCs Isolated PBMCs were divided into 1 × 106 cells per tube (each 100 μ L of PBS) and incubated with surface monoclonal antibodies against CD3, CD4, CD8, and CD19 conjugated with the fluorochromes allophycocyanin (APC), peridinin chlorophyll protein (Per-CP), and phycoerythrin-Cy7 (PE-Cy7) (all from BD Pharmingen, San Diego, CA, USA) at a concentration of 20 μ L/1 × 106 cells, in darkness at room temperature for 30 min. .. The cells were then fixed and permeabilized using an intracellular TLR staining kit according to the producer's protocol (Imgenex, San Diego, CA, USA).

    Article Title: Exposure to different early-life stress experiences results in differentially altered DNA methylation in the brain and immune system
    Article Snippet: .. 2.10 Human PBMCs isolation and DNA purificationBlood (5 ml) was drawn on all participants, collected in EDTA vacutainer (BD, Toronto, Ontario, Canada), and PBMCs and DNA were isolated within few hours from collection as described for mouse studies. .. Global methylation quantification was performed by ELISA using the Epigentek Methylflash Methylated DNA Quantification Kit methyas described for mouse studies.

    Article Title: Impact of genetic risk loci for multiple sclerosis on expression of proximal genes in patients.
    Article Snippet: .. Preparation of PBMCs, CD4þ and CD8þ T cellsWe isolated PBMCs immediately from blood samples taken with sodium citrate-containing cell preparation tubes (BD VacutainerTM CPTTM Tube, Becton-Dickinson, Franklin Lakes, NJ), in the screening phase, or using a standard Ficoll (GE Healthcare, Little Chalfont, UK) procedure, in the validation phase. .. We separated the cells with density gradient centrifugation, collected them from the interphase, washed twice in Dubelcco’s phosphate buffered saline (PBS) and either froze as a pellet at 80 C for subsequent RNA extraction, or prepared the cells for sorting.

    Article Title: SLE non-coding genetic risk variant determines the epigenetic dysfunction of an immune cell specific enhancer that controls disease-critical microRNA expression
    Article Snippet: .. PBMCs isolation Whole blood was collected from healthy human donors or SLE patient donors in sodium heparinized vacutainer tubes (Becton Dickinson, USA) with approval by the Committee on Human Research of Renji Hospital, Shanghai Jiao Tong University. ..

    Article Title: The effect of moderate weight loss, with or without (1, 3)(1, 6)-β-glucan addition, on subcutaneous adipose tissue inflammatory gene expression in young subjects with uncomplicated obesity
    Article Snippet: AT biopsy Subcutaneous AT biopsy was obtained from the umbilical region using biopsy needle and collected to 1 ml of RNA stabilization reagent (Allprotect Tissue Reagent, Qiagen GmbH, Hilden, Germany), as described [ – ]. .. PBMC isolation PBMC were isolated from fresh blood using 10 mL BD Vacutainer® CPT™ Cell Preparation Tube (Becton Dickinson AG, NJ, USA) with sodium citrate, following the instructions of the manufacturer. ..

    Incubation:

    Article Title: Biologically driven cut-off definition of lymphocyte ratios in metastatic breast cancer and association with exosomal subpopulations and prognosis
    Article Snippet: Exosome internalization experiments To compare the ability of CD14 + monocytes and CD3 + lymphocytes to internalize plasma-derived exosomes, 1.5 × 1010 exosomes plasma-derived were resuspended in PBS and stained with 5 μM DiD for 30 minutes at 37 °C. .. 2 × 105 isolated PBMCs were then incubated with DiD-labelled vesicles for 5 and 24 hours, respectively, and subsequently stained for CD45, CD3 and CD14 (Becton Dickinson) and analyzed by flow cytometry (FACSCanto II, Becton Dickinson) by gating either on CD14 + or CD3 + PBMCs. ..

    Article Title: Expression of Toll-Like Receptors 3, 7, and 9 in Peripheral Blood Mononuclear Cells from Patients with Systemic Lupus Erythematosus
    Article Snippet: .. Assessment of TLR Expression in PBMCs Isolated PBMCs were divided into 1 × 106 cells per tube (each 100 μ L of PBS) and incubated with surface monoclonal antibodies against CD3, CD4, CD8, and CD19 conjugated with the fluorochromes allophycocyanin (APC), peridinin chlorophyll protein (Per-CP), and phycoerythrin-Cy7 (PE-Cy7) (all from BD Pharmingen, San Diego, CA, USA) at a concentration of 20 μ L/1 × 106 cells, in darkness at room temperature for 30 min. .. The cells were then fixed and permeabilized using an intracellular TLR staining kit according to the producer's protocol (Imgenex, San Diego, CA, USA).

    Staining:

    Article Title: Biologically driven cut-off definition of lymphocyte ratios in metastatic breast cancer and association with exosomal subpopulations and prognosis
    Article Snippet: Exosome internalization experiments To compare the ability of CD14 + monocytes and CD3 + lymphocytes to internalize plasma-derived exosomes, 1.5 × 1010 exosomes plasma-derived were resuspended in PBS and stained with 5 μM DiD for 30 minutes at 37 °C. .. 2 × 105 isolated PBMCs were then incubated with DiD-labelled vesicles for 5 and 24 hours, respectively, and subsequently stained for CD45, CD3 and CD14 (Becton Dickinson) and analyzed by flow cytometry (FACSCanto II, Becton Dickinson) by gating either on CD14 + or CD3 + PBMCs. ..

    Flow Cytometry:

    Article Title: Biologically driven cut-off definition of lymphocyte ratios in metastatic breast cancer and association with exosomal subpopulations and prognosis
    Article Snippet: Exosome internalization experiments To compare the ability of CD14 + monocytes and CD3 + lymphocytes to internalize plasma-derived exosomes, 1.5 × 1010 exosomes plasma-derived were resuspended in PBS and stained with 5 μM DiD for 30 minutes at 37 °C. .. 2 × 105 isolated PBMCs were then incubated with DiD-labelled vesicles for 5 and 24 hours, respectively, and subsequently stained for CD45, CD3 and CD14 (Becton Dickinson) and analyzed by flow cytometry (FACSCanto II, Becton Dickinson) by gating either on CD14 + or CD3 + PBMCs. ..

    Expressing:

    Article Title: Expression of Toll-Like Receptors 3, 7, and 9 in Peripheral Blood Mononuclear Cells from Patients with Systemic Lupus Erythematosus
    Article Snippet: .. Assessment of TLR Expression in PBMCs Isolated PBMCs were divided into 1 × 106 cells per tube (each 100 μ L of PBS) and incubated with surface monoclonal antibodies against CD3, CD4, CD8, and CD19 conjugated with the fluorochromes allophycocyanin (APC), peridinin chlorophyll protein (Per-CP), and phycoerythrin-Cy7 (PE-Cy7) (all from BD Pharmingen, San Diego, CA, USA) at a concentration of 20 μ L/1 × 106 cells, in darkness at room temperature for 30 min. .. The cells were then fixed and permeabilized using an intracellular TLR staining kit according to the producer's protocol (Imgenex, San Diego, CA, USA).

    Concentration Assay:

    Article Title: Expression of Toll-Like Receptors 3, 7, and 9 in Peripheral Blood Mononuclear Cells from Patients with Systemic Lupus Erythematosus
    Article Snippet: .. Assessment of TLR Expression in PBMCs Isolated PBMCs were divided into 1 × 106 cells per tube (each 100 μ L of PBS) and incubated with surface monoclonal antibodies against CD3, CD4, CD8, and CD19 conjugated with the fluorochromes allophycocyanin (APC), peridinin chlorophyll protein (Per-CP), and phycoerythrin-Cy7 (PE-Cy7) (all from BD Pharmingen, San Diego, CA, USA) at a concentration of 20 μ L/1 × 106 cells, in darkness at room temperature for 30 min. .. The cells were then fixed and permeabilized using an intracellular TLR staining kit according to the producer's protocol (Imgenex, San Diego, CA, USA).

    Cycling Probe Technology:

    Article Title: The effect of moderate weight loss, with or without (1, 3)(1, 6)-β-glucan addition, on subcutaneous adipose tissue inflammatory gene expression in young subjects with uncomplicated obesity
    Article Snippet: AT biopsy Subcutaneous AT biopsy was obtained from the umbilical region using biopsy needle and collected to 1 ml of RNA stabilization reagent (Allprotect Tissue Reagent, Qiagen GmbH, Hilden, Germany), as described [ – ]. .. PBMC isolation PBMC were isolated from fresh blood using 10 mL BD Vacutainer® CPT™ Cell Preparation Tube (Becton Dickinson AG, NJ, USA) with sodium citrate, following the instructions of the manufacturer. ..

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    • peripheral blood mononuclear cell pbmc isolation  (Becton Dickinson)
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    Becton Dickinson peripheral blood mononuclear cell pbmc isolation
    mRNA level in <t>PBMCs</t> measured over the duration of intervention period for (A) SEPW1, (B) SEPS1, (C) SEPR in the placebo and Se-yeast groups. Values are means ± SEM relative to baseline, week 0 expression. GUSB was used as reference gene for normalisation. White bars = placebo group (n = 14 to 20); light grey bars = 50 µg/day Se-yeast (n = 11 to 15); mid grey bars = 100 µg/day Se-yeast (n = 10 to 19); black bars = 200 µg/day Se-yeast (n = 14 to 18). The variation in sample number (n) between time points for each treatment is due to insufficient <t>RNA</t> at some sampling time points and missing time course sample data for some of the target genes. For the SEPS1 gene expression data set: placebo group n = 20 at all time points; 50 µg/day group n = 14 at wks 0, 6 and 12 (n = 13 at wks 10, 11); 100 µg/day group n = 19 at wks 0, 6, 10 (n = 18 at wks 11, 12); 200 µg/day group n = 18 at wks 0, 6, 12 (n = 17 wk 10, n = 15 wk 11). Data were analysed using mixed-effects models and statistically significant differences are indicated on the figure.
    Peripheral Blood Mononuclear Cell Pbmc Isolation, 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/peripheral blood mononuclear cell pbmc isolation/product/Becton Dickinson
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    peripheral blood mononuclear cell pbmc isolation - by Bioz Stars, 2021-03
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    pbmc isolation  (Becton Dickinson)
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    Becton Dickinson pbmc isolation
    ( a ) p24 viral antigen levels 10 days post-infection in the supernatant of <t>HIV-1</t> infected PBMCs treated with TIZ (10 μg/mL) or RM-4848 (10 μg/mL). Results obtained with <t>PBMC</t> of 20 different healthy donors are shown. Median values and statistical significance is presented. ( b ) Inhibition (%) of viral replication in PBMC of 20 different healthy donors infected in vitro with HIV-1 in the presence of TIZ- (10 μg/mL) or RM-4848- (10 μg/mL). Median values and statistical significance is presented. ( c ) p24 viral antigen levels in PBMCs infected with HIV-1 in the presence of RM-4848 throughout infection (RM-4848)(10 μg/mL) or in medium alone (no treatment). In addition to these two situations, results of 3 other experimental conditions are shown: 1) PBMCs treated with RM-4848 before being infected (pre-infection); 2) RM-4848 present during the 24 hours infection period (during infection) and then washed away; 3) RM-4848 added 24 hours after infection (post infection). Mean values ± standard errors are shown.
    Pbmc Isolation, 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/pbmc isolation/product/Becton Dickinson
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    pbmc isolation - by Bioz Stars, 2021-03
    86/100 stars
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    pbmc isolation peripheral blood mononuclear cells pbmcs  (Becton Dickinson)
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    Becton Dickinson pbmc isolation peripheral blood mononuclear cells pbmcs
    Cellular response against IFN-γ inducer peptides by multiparous and hyper-vaccinated sows. The IFNγ-SC frequencies between groups of sows against the pooled peptides from nsp and structural proteins of PRRSV were determined by ELISPOT IFN-γ assays of <t>PBMCs.</t> No statistical significance was observed in the IFNγ-SC frequency between sow groups. Each bar represents the mean ± standard deviation. White, black and gray bars represent the groups of sows with one, three and six parities, respectively.
    Pbmc Isolation Peripheral Blood Mononuclear Cells Pbmcs, 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/pbmc isolation peripheral blood mononuclear cells pbmcs/product/Becton Dickinson
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    pbmc isolation peripheral blood mononuclear cells pbmcs - by Bioz Stars, 2021-03
    86/100 stars
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    mRNA level in PBMCs measured over the duration of intervention period for (A) SEPW1, (B) SEPS1, (C) SEPR in the placebo and Se-yeast groups. Values are means ± SEM relative to baseline, week 0 expression. GUSB was used as reference gene for normalisation. White bars = placebo group (n = 14 to 20); light grey bars = 50 µg/day Se-yeast (n = 11 to 15); mid grey bars = 100 µg/day Se-yeast (n = 10 to 19); black bars = 200 µg/day Se-yeast (n = 14 to 18). The variation in sample number (n) between time points for each treatment is due to insufficient RNA at some sampling time points and missing time course sample data for some of the target genes. For the SEPS1 gene expression data set: placebo group n = 20 at all time points; 50 µg/day group n = 14 at wks 0, 6 and 12 (n = 13 at wks 10, 11); 100 µg/day group n = 19 at wks 0, 6, 10 (n = 18 at wks 11, 12); 200 µg/day group n = 18 at wks 0, 6, 12 (n = 17 wk 10, n = 15 wk 11). Data were analysed using mixed-effects models and statistically significant differences are indicated on the figure.

    Journal: PLoS ONE

    Article Title: Effects of Selenium Supplementation on Selenoprotein Gene Expression and Response to Influenza Vaccine Challenge: A Randomised Controlled Trial

    doi: 10.1371/journal.pone.0014771

    Figure Lengend Snippet: mRNA level in PBMCs measured over the duration of intervention period for (A) SEPW1, (B) SEPS1, (C) SEPR in the placebo and Se-yeast groups. Values are means ± SEM relative to baseline, week 0 expression. GUSB was used as reference gene for normalisation. White bars = placebo group (n = 14 to 20); light grey bars = 50 µg/day Se-yeast (n = 11 to 15); mid grey bars = 100 µg/day Se-yeast (n = 10 to 19); black bars = 200 µg/day Se-yeast (n = 14 to 18). The variation in sample number (n) between time points for each treatment is due to insufficient RNA at some sampling time points and missing time course sample data for some of the target genes. For the SEPS1 gene expression data set: placebo group n = 20 at all time points; 50 µg/day group n = 14 at wks 0, 6 and 12 (n = 13 at wks 10, 11); 100 µg/day group n = 19 at wks 0, 6, 10 (n = 18 at wks 11, 12); 200 µg/day group n = 18 at wks 0, 6, 12 (n = 17 wk 10, n = 15 wk 11). Data were analysed using mixed-effects models and statistically significant differences are indicated on the figure.

    Article Snippet: Peripheral blood mononuclear cell (PBMC) isolation and preparation of total RNA for quantitative real time RT-PCR Approximately 8 ml of whole blood was collected in a BD Vacutainer™ CPT™ tube (BD Medical, Cowley, UK, http://www.bd.com/ ).

    Techniques: Expressing, Sampling

    ( a ) p24 viral antigen levels 10 days post-infection in the supernatant of HIV-1 infected PBMCs treated with TIZ (10 μg/mL) or RM-4848 (10 μg/mL). Results obtained with PBMC of 20 different healthy donors are shown. Median values and statistical significance is presented. ( b ) Inhibition (%) of viral replication in PBMC of 20 different healthy donors infected in vitro with HIV-1 in the presence of TIZ- (10 μg/mL) or RM-4848- (10 μg/mL). Median values and statistical significance is presented. ( c ) p24 viral antigen levels in PBMCs infected with HIV-1 in the presence of RM-4848 throughout infection (RM-4848)(10 μg/mL) or in medium alone (no treatment). In addition to these two situations, results of 3 other experimental conditions are shown: 1) PBMCs treated with RM-4848 before being infected (pre-infection); 2) RM-4848 present during the 24 hours infection period (during infection) and then washed away; 3) RM-4848 added 24 hours after infection (post infection). Mean values ± standard errors are shown.

    Journal: Scientific Reports

    Article Title: Thiazolides Elicit Anti-Viral Innate Immunity and Reduce HIV Replication

    doi: 10.1038/srep27148

    Figure Lengend Snippet: ( a ) p24 viral antigen levels 10 days post-infection in the supernatant of HIV-1 infected PBMCs treated with TIZ (10 μg/mL) or RM-4848 (10 μg/mL). Results obtained with PBMC of 20 different healthy donors are shown. Median values and statistical significance is presented. ( b ) Inhibition (%) of viral replication in PBMC of 20 different healthy donors infected in vitro with HIV-1 in the presence of TIZ- (10 μg/mL) or RM-4848- (10 μg/mL). Median values and statistical significance is presented. ( c ) p24 viral antigen levels in PBMCs infected with HIV-1 in the presence of RM-4848 throughout infection (RM-4848)(10 μg/mL) or in medium alone (no treatment). In addition to these two situations, results of 3 other experimental conditions are shown: 1) PBMCs treated with RM-4848 before being infected (pre-infection); 2) RM-4848 present during the 24 hours infection period (during infection) and then washed away; 3) RM-4848 added 24 hours after infection (post infection). Mean values ± standard errors are shown.

    Article Snippet: PBMC isolation and HIV-1 infection Whole blood was collected from 20 healthy volunteers by venipuncture in Vacutainer tubes containing EDTA (Becton Dickinson, Franklin Lakes, NJ), and PBMCs were separated on lymphocyte separation medium (Organon Teknica, Malvern, PA).

    Techniques: Infection, Inhibition, In Vitro

    Cytokines (IL-2, IFNγ) and chemokines (MCP-1, MIP-1α, MIP-1β, RANTES) production (pg/ml) by PBMCs of healthy donors. PBMCs were either uninfected (white bars) or HIV-1-infected (grey bars) and were cultured in medium alone (no drug), or in medium containing either TIZ- (10 μg/mL) ( a ) or RM-4848- (10 μg/mL) ( b ). Mean values 7 days post-infection ± standard errors and statistical differences are shown.

    Journal: Scientific Reports

    Article Title: Thiazolides Elicit Anti-Viral Innate Immunity and Reduce HIV Replication

    doi: 10.1038/srep27148

    Figure Lengend Snippet: Cytokines (IL-2, IFNγ) and chemokines (MCP-1, MIP-1α, MIP-1β, RANTES) production (pg/ml) by PBMCs of healthy donors. PBMCs were either uninfected (white bars) or HIV-1-infected (grey bars) and were cultured in medium alone (no drug), or in medium containing either TIZ- (10 μg/mL) ( a ) or RM-4848- (10 μg/mL) ( b ). Mean values 7 days post-infection ± standard errors and statistical differences are shown.

    Article Snippet: PBMC isolation and HIV-1 infection Whole blood was collected from 20 healthy volunteers by venipuncture in Vacutainer tubes containing EDTA (Becton Dickinson, Franklin Lakes, NJ), and PBMCs were separated on lymphocyte separation medium (Organon Teknica, Malvern, PA).

    Techniques: Infection, Cell Culture

    mRNA expression of 84 genes that are part of the HIV-1 life cycle and known to directly obstacle HIV infectivity has been assessed by real-time quantitative RT-PCR, calculated relative to five housekeeping genes and shown as fold-change expression from the untreated sample. Gene expression (nfold) is shown as a color scale from green to red (−2 to +9) (MEV multiple experiment viewer software). Results were obtained in HIV1-infected PBMCs at day 7 post infection, cultured in medium containing TIZ (10 μg/mL) or RM-4848 (10 μg/mL). Only targets showing > 2-fold modulation are considered significant and are shown in tab.

    Journal: Scientific Reports

    Article Title: Thiazolides Elicit Anti-Viral Innate Immunity and Reduce HIV Replication

    doi: 10.1038/srep27148

    Figure Lengend Snippet: mRNA expression of 84 genes that are part of the HIV-1 life cycle and known to directly obstacle HIV infectivity has been assessed by real-time quantitative RT-PCR, calculated relative to five housekeeping genes and shown as fold-change expression from the untreated sample. Gene expression (nfold) is shown as a color scale from green to red (−2 to +9) (MEV multiple experiment viewer software). Results were obtained in HIV1-infected PBMCs at day 7 post infection, cultured in medium containing TIZ (10 μg/mL) or RM-4848 (10 μg/mL). Only targets showing > 2-fold modulation are considered significant and are shown in tab.

    Article Snippet: PBMC isolation and HIV-1 infection Whole blood was collected from 20 healthy volunteers by venipuncture in Vacutainer tubes containing EDTA (Becton Dickinson, Franklin Lakes, NJ), and PBMCs were separated on lymphocyte separation medium (Organon Teknica, Malvern, PA).

    Techniques: Expressing, Infection, Quantitative RT-PCR, Software, Cell Culture

    mRNA expression of genes involved in cholesterol metabolism and efflux in PBMCs of healthy donors. ( a ) Uninfected PBMCs cultured in medium containing TIZ (10 μg/mL); ( b ) Uninfected PBMCs cultured in medium containing RM-4848 (10 μg/mL); ( c ) HIV-1–infected PBMCs (day 7 post infection) cultured in medium containing TIZ (10 μg/mL); ( d ) HIV-1–infected PBMCs (day 7 post infection) cultured in medium containing RM-4848 (10 μg/mL). Gene expression genes is assessed by quantitative RT-PCR and shown as fold-change expression from the untreated sample. Only targets showing > 2-fold modulation are considered significant. Mean values ± S.D. and p values are indicated.

    Journal: Scientific Reports

    Article Title: Thiazolides Elicit Anti-Viral Innate Immunity and Reduce HIV Replication

    doi: 10.1038/srep27148

    Figure Lengend Snippet: mRNA expression of genes involved in cholesterol metabolism and efflux in PBMCs of healthy donors. ( a ) Uninfected PBMCs cultured in medium containing TIZ (10 μg/mL); ( b ) Uninfected PBMCs cultured in medium containing RM-4848 (10 μg/mL); ( c ) HIV-1–infected PBMCs (day 7 post infection) cultured in medium containing TIZ (10 μg/mL); ( d ) HIV-1–infected PBMCs (day 7 post infection) cultured in medium containing RM-4848 (10 μg/mL). Gene expression genes is assessed by quantitative RT-PCR and shown as fold-change expression from the untreated sample. Only targets showing > 2-fold modulation are considered significant. Mean values ± S.D. and p values are indicated.

    Article Snippet: PBMC isolation and HIV-1 infection Whole blood was collected from 20 healthy volunteers by venipuncture in Vacutainer tubes containing EDTA (Becton Dickinson, Franklin Lakes, NJ), and PBMCs were separated on lymphocyte separation medium (Organon Teknica, Malvern, PA).

    Techniques: Expressing, Cell Culture, Infection, Quantitative RT-PCR

    p24 viral antigen concentration in HIV-1 - infected PBMCs cultured in medium alone (no drug) or in the presence of TO-901317 (1 μM), an LXR agonist. Results were obtained with PBMCs of 20 different healthy donors 10 days post-infection. Mean values ± standard errors and statistical significance are shown.

    Journal: Scientific Reports

    Article Title: Thiazolides Elicit Anti-Viral Innate Immunity and Reduce HIV Replication

    doi: 10.1038/srep27148

    Figure Lengend Snippet: p24 viral antigen concentration in HIV-1 - infected PBMCs cultured in medium alone (no drug) or in the presence of TO-901317 (1 μM), an LXR agonist. Results were obtained with PBMCs of 20 different healthy donors 10 days post-infection. Mean values ± standard errors and statistical significance are shown.

    Article Snippet: PBMC isolation and HIV-1 infection Whole blood was collected from 20 healthy volunteers by venipuncture in Vacutainer tubes containing EDTA (Becton Dickinson, Franklin Lakes, NJ), and PBMCs were separated on lymphocyte separation medium (Organon Teknica, Malvern, PA).

    Techniques: Concentration Assay, Infection, Cell Culture

    Box-and-whisker plot of HAVCR2 expression depending on rs4704846 genotype. Data derive from PBMCs from 40 healthy volunteers uninfected or infected with HIV-1. HAVCR2 transcript levels are log-transformed and shown in standard box-and-whisker plot representation (thick line: median; box: quartiles; whiskers: 1.5 × interquartile range); p values are calculated using the Student's t -test.

    Journal: PLoS ONE

    Article Title: A Regulatory Polymorphism in HAVCR2 Modulates Susceptibility to HIV-1 Infection

    doi: 10.1371/journal.pone.0106442

    Figure Lengend Snippet: Box-and-whisker plot of HAVCR2 expression depending on rs4704846 genotype. Data derive from PBMCs from 40 healthy volunteers uninfected or infected with HIV-1. HAVCR2 transcript levels are log-transformed and shown in standard box-and-whisker plot representation (thick line: median; box: quartiles; whiskers: 1.5 × interquartile range); p values are calculated using the Student's t -test.

    Article Snippet: PBMC isolation, HIV-1 infection, and transcript quantification Whole blood was collected from 40 healthy volunteers by venipuncture in Vacutainer tubes containing EDTA (Becton Dickinson), and PBMCs were separated on lymphocyte separation medium (Organon Teknica, Malvern, PA).

    Techniques: Whisker Assay, Expressing, Infection, Transformation Assay

    Cellular response against IFN-γ inducer peptides by multiparous and hyper-vaccinated sows. The IFNγ-SC frequencies between groups of sows against the pooled peptides from nsp and structural proteins of PRRSV were determined by ELISPOT IFN-γ assays of PBMCs. No statistical significance was observed in the IFNγ-SC frequency between sow groups. Each bar represents the mean ± standard deviation. White, black and gray bars represent the groups of sows with one, three and six parities, respectively.

    Journal: Vaccines

    Article Title: Immune Response of Multiparous Hyper-Immunized Sows against Peptides from Non-Structural and Structural Proteins of PRRSV

    doi: 10.3390/vaccines3040973

    Figure Lengend Snippet: Cellular response against IFN-γ inducer peptides by multiparous and hyper-vaccinated sows. The IFNγ-SC frequencies between groups of sows against the pooled peptides from nsp and structural proteins of PRRSV were determined by ELISPOT IFN-γ assays of PBMCs. No statistical significance was observed in the IFNγ-SC frequency between sow groups. Each bar represents the mean ± standard deviation. White, black and gray bars represent the groups of sows with one, three and six parities, respectively.

    Article Snippet: PBMC Isolation Peripheral blood mononuclear cells (PBMCs) from hyper-immunized sows were collected into heparin-coated tubes (Becton-Dickinson, BD, Franklin Lakes, NY, USA).

    Techniques: Enzyme-linked Immunospot, Standard Deviation