Anti cd3 anti cd28 human t activator dynabeads

Manufactured by Thermo Fisher Scientific

The Anti-CD3/anti-CD28 human T-Activator Dynabeads are uniform, superparamagnetic beads coated with monoclonal antibodies specific for the CD3 and CD28 receptors on human T cells. These beads are designed to activate and expand human T cells in vitro.

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Lab products found in correlation

Ficoll paque plus, by GE Healthcare (3 mentions) Memory cd4 t cell enrichment kits, by STEMCELL (2 mentions) Qiazol lysis reagent, by Qiagen (2 mentions) Anti cd3 apc h7, by BD (1 mentions) Anti cd4 pe cy7, by BD (1 mentions) Anti cd4 microbeads, by Miltenyi Biotec (1 mentions) Lipopolysaccharides (lps), by Merck Group (1 mentions) Cpg odn 2006, by InvivoGen (1 mentions) Anti cd14 fitc, by Miltenyi Biotec (1 mentions) Anti cd11b apc, by Miltenyi Biotec (1 mentions) Anti cd8 pe, by Miltenyi Biotec (1 mentions)

Spelling variants of this product

CD3/CD28 Dynabeads (277 citations) Anti-CD3/CD28 Dynabeads (179 citations) Anti-CD3/anti-CD28 Dynabeads (63 citations) Anti-CD3/CD28 (45 citations) Anti-human CD3 (35 citations) Dynabeads CD3 (25 citations) Anti-human CD3/CD28 Dynabeads (20 citations) Human T-Activator CD3/CD28 (12 citations) Anti-human CD28 (6 citations) Anti-CD3/CD28 Human T-Activator Dynabeads (2 citations)

5 protocols using anti cd3 anti cd28 human t activator dynabeads

Isolation and Stimulation of Naive and Memory CD4+ T Cells

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Blood samples were obtained from 119 healthy individuals of British ancestry. Of these, 67 were male (53.7%) and 52 female (56.3%), and the mean age of the cohort was 47 years (sd = 15.61 years) (Supplementary Figure 1a). Human biological samples were sourced ethically and their research use was in accord with the terms of informed consent under an IRB/EC approved protocol (15/NW/0282). Peripheral blood mononuclear cells (PBMCs) were isolated using Ficoll-Paque PLUS (GE healthcare, Buckingham, UK) density gradient centrifugation. Naïve (CD25- CD45RA+ CD45RO-) and memory (CD25- CD45RA- CD45RO+) CD4+ T cells were isolated from the PBMC fraction using EasySep® naïve CD4+ T cell isolation kits and memory CD4+ T cell enrichment kits (StemCell Technologies, Meylan, France) according to the manufacturer's instructions. Naive and memory T cells were then stimulated with anti-CD3/anti-CD28 human T-Activator Dynabeads® (Invitrogen) at a 1:2 beads-to-cells ratio. Cells were harvested after 16 hours, 40 hours, and 5 days of stimulation. In addition, unstimulated cells kept in culture without any beads for 16 hours were used as a negative control (i.e. zero hours of activation).

Soskic B., Cano-Gamez E., Smyth D.J., Ambridge K., Ke Z., Matte J.C., Bossini-Castillo L., Kaplanis J., Ramirez-Navarro L., Lorenc A., Nakic N., Esparza-Gordillo J., Rowan W., Wille D., Tough D.F., Bronson P.G, & Trynka G. (2022). Immune disease risk variants regulate gene expression dynamics during CD4+ T cell activation. Nature Genetics, 54(6), 817-826.

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Isolation and Stimulation of Naive and Memory CD4+ T Cells

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Blood samples were obtained from 119 healthy individuals of British ancestry. Of these, 67 were male (53.7%) and 52 female (56.3%), and the mean age of the cohort was 47 years (standard deviation = 15.61 years) (Supplementary Fig. 1a). Human biological samples were sourced ethically, and their research use was in accord with the terms of informed consent under an institutional review board/ethics committee-approved protocol (15/NW/0282).
Peripheral blood mononuclear cells (PBMCs) were isolated using Ficoll-Paque PLUS (GE Healthcare) density gradient centrifugation. Naive (CD25⁻ CD45RA⁺ CD45RO⁻) and memory (CD25⁻ CD45RA⁻ CD45RO⁺) CD4⁺ T cells were isolated from the PBMC fraction using EasySep naive CD4⁺ T cell isolation kits and memory CD4⁺ T cell enrichment kits (StemCell Technologies) according to the manufacturer’s instructions. Naive and memory T cells were then stimulated with anti-CD3/anti-CD28 human T-Activator Dynabeads (Invitrogen) at a 1:2 beads-to-cells ratio. Cells were harvested after 16 h, 40 h and 5 d of stimulation. In addition, unstimulated cells kept in culture without any beads for 16 h were used as a negative control (i.e., 0 h of activation).

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CD4+ T-cell Isolation and Activation

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Blood was drawn into Sodium Heparin tubes (APP Pharmaceuticals, NDC 63323-540-11); CD4+ cells were isolated from whole blood using anti-CD4 Microbeads (Miltenyi Biotec) and MS column purification as per manufacturer’s instructions. A small portion of the cell isolate was set aside for flow cytometry using anti CD3 APC-H7 and anti CD4 PE-Cy7 (BD Pharmingen) to assess purity. Isolated cells were re-suspended at 1 million/ml in RPMI containing 10% FBS and either immediately lysed in 0.7 ml QIAzol lysis reagent (Qiagen) or incubated for 4 hours at 37°C with 25 µl/ml anti-CD3/anti-CD28 Human T-Activator Dynabeads (Invitrogen) prior to lysis. Lysates were stored at −80°C until processing for RNAseq.

Felts S.J., Van Keulen V.P., Scheid A.D., Allen K.S., Bradshaw R.K., Jen J., Peikert T., Middha S., Zhang Y., Block M.S., Markovic S.N, & Pease L.R. (2015). Gene Expression Patterns in CD4+ Peripheral Blood Cells in Healthy Subjects and Stage IV Melanoma Patients. Cancer immunology, immunotherapy : CII, 64(11), 1437-1447.

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Isolation and Stimulation of Human T Cells

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Blood samples were obtained from six individuals for the bulk assays (naïve and memory T cells were isolated from three independent individuals, respectively) and from four additional individuals for single-cell RNA-seq. All individuals were healthy males of 56.4 years of age on average (sd = 12.41 years). Human biological samples were sourced ethically and their research use was in accord with the terms of the informed consents under an IRB/EC approved protocol (15/NW/0282). Peripheral blood mononuclear cells (PBMCs) were isolated using Ficoll-Paque PLUS (GE healthcare, Buckingham, UK) density gradient centrifugation. Naïve (CD25− CD45RA + CD45RO−) and memory (CD25− CD45RA− CD45RO+) CD4⁺ T cells were isolated from PBMCs using EasySep® naïve CD4⁺ T cell isolation kit and memory CD4⁺ T cell enrichment kit (StemCell Technologies, Meylan, France) according to the manufacturer’s instructions. T cells were then stimulated with anti-CD3/anti-CD28 human T-Activator Dynabeads® (Invitrogen) at a 1:2 ratio of beads to T cells. Cytokines were added at the same time as the stimulus (see Supplementary Data 1 for a full list of the cytokines used with product details and exact concentrations). Cells were harvested after 16 h and 5 days of stimulation.

Cano-Gamez E., Soskic B., Roumeliotis T.I., So E., Smyth D.J., Baldrighi M., Willé D., Nakic N., Esparza-Gordillo J., Larminie C.G., Bronson P.G., Tough D.F., Rowan W.C., Choudhary J.S, & Trynka G. (2020). Single-cell transcriptomics identifies an effectorness gradient shaping the response of CD4+ T cells to cytokines. Nature Communications, 11, 1801.

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Isolation and Activation of Immune Cells

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Blood was drawn into sodium heparin tubes (APP Pharmaceuticals, NDC 63323-540-11). CD4+, CD8+, and CD14+ cells were isolated from whole blood using anti-CD4/CD8/CD14 microbeads (130-090-877/130-090-878/130-090-879, Miltenyi Biotec) and Automatics purification as per the manufacturer’s instructions. A small portion of the cell isolate was set aside for flow cytometry using anti-CD3 APC-H7, anti-CD4 PE-Cy7, anti-CD11b-APC, anti-CD8 PE, anti CD14 – FITC (Miltenyi Biotec) to assess the purity. All antibodies were purchased from BD Pharmingen unless otherwise indicated. Isolated cells were re-suspended at 1 million/ml in RPMI containing 10% FBS and either immediately lysed in 0.7 ml QIAzol lysis reagent (Qiagen) or incubated for 4 h at 37 °C with 25 μl/ml anti-CD3/antiCD28 Human T-Activator Dynabeads (111.32D, Invitrogen) for CD4 and CD8 cells or 1 μg/ml pI:C (528906, Calbiochem), LPS (L439, Sigma) CPG ODN2006, Invivogen) and PGN (Sigma, 77140) for CD14 cells. Lysates were stored at −80 °C until processing for RNAseq.

Wang L., Felts S.J., Van Keulen V.P., Scheid A.D., Block M.S., Markovic S.N., Pease L.R, & Zhang Y. (2018). Integrative genome-wide analysis of long noncoding RNAs in diverse immune cell types of melanoma patients. Cancer research, 78(15), 4411-4423.

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