The following Abs and isotype controls were purchased from e-Biosciences: PE-conjugated anti–IFN-γ and rat IgG2a; PE-conjugated anti–IL-5; PECy5-labeled anti-CD4; anti-CD80 (16-10A1 ) and anti-CD83 (Michel-17); rat IgG1, IgG2a, and IgG2b; PE-conjugated anti-CD11c (N418); hamster IgG; PE-conjugated anti–IL-17A (eBio17B7); and rat IgG2a. For intracellular staining, cell suspensions from cervical lymph nodes (cLNs) were stimulated with anti-CD3 and anti-CD28 Abs (eBioscience) for 48 h, followed by PMA (50 ng/ml), ionomycin (1 µg/ml), and brefeldin A (10 µg/ml) for 6 h. Cells were washed, incubated with FcR block (1 µg/ml; eBiosciences), and stained for CD4. Cells were fixed with 2% paraformaldehyde, permeabilized with 0.1% saponin (Sigma), and stained for IL-5, IL-17 and IFN-γ. Cells were analyzed on a FACS Calibur (BD Biosciences) with FCS Express software (DeNovo Software).
Anti cd3 and anti cd28 abs
Manufactured by Thermo Fisher Scientific
Sourced in United States
Anti-CD3 and anti-CD28 Abs are laboratory reagents that bind to specific cell surface markers, CD3 and CD28, on T cells. These antibodies are commonly used to activate and stimulate T cell proliferation in cell culture experiments and research applications.
Automatically generated - may contain errors
Lab products found in correlation
Pe conjugated anti il 17a, by Thermo Fisher Scientific (1 mentions)
Rat igg2a, by Thermo Fisher Scientific (1 mentions)
Fcs express software, by De Novo Software (1 mentions)
Fcr block, by Thermo Fisher Scientific (1 mentions)
Pe conjugated anti il 5, by Thermo Fisher Scientific (1 mentions)
Saponin, by Merck Group (1 mentions)
Facscalibur, by BD (1 mentions)
Anti il 4 ab, by R&D Systems (1 mentions)
Anti il 21 ab, by R&D Systems (1 mentions)
Il 12, by Fujifilm (1 mentions)
Interleukin 6 (il 6), by Thermo Fisher Scientific (1 mentions)
3 protocols using anti cd3 and anti cd28 abs
1
Multiparametric Flow Cytometry Analysis
2
Th17 cell differentiation from naive T cells
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Naive T cells from young mice were stimulated with plate-bound anti-CD3 and anti-CD28 Abs (both eBioscience) in the presence of IL-12 (8 ng ml−1; Wako) at 2.5 × 105 cells ml−1 for 5 days. IL-6 (10 ng ml−1; Peprotech), IL-21 (20 ng ml−1; Wako), anti-IL-4 Ab (10 μg ml−1) and/or anti-IL-21 Ab (10 μg ml−1; R&D Systems) was added to the culture.
3
Exosome-Mediated T Cell Proliferation Assay
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CD4+ T cells isolated from PBMC of normal donors as described above were labelled with 1.5 µM CSFE (Cell Trace, Life Technologies) in 0.1% BSA in PBS (w/v) for 10 min at 37°C. The staining was quenched with an equal volume of exosome-depleted FBS (13 (link)). CFSE-labelled T cells (105/well) were incubated with plate-bound anti-CD3 and anti-CD28 Abs (3 µg/mL; eBioscience, San Diego, CA, USA) for 24 h and following activation, were co-incubated with exosomes (50 µL aliquots of SEC fractions #4) for 4 days at 37°C. Proliferation of T cells was measured on day 5 by flow cytometry, and the data were analysed by Modfit (Verity Software House, Topsham, ME, USA). The percent suppression of proliferation in co-cultures with exosomes was calculated as described by Strauss et al. (15 (link)) and compared to control T cells incubated alone.
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