Memory B cells were differentiated in vitro into plasmablasts and plasma cells, following the protocol described in Jourdan et al. (24 (link)). Briefly, CD27+ Memory B cells were first purified as bulk B cells by negative selection from PBMCs by magnetic isolation followed by positive selection for CD27+ cells (Stemcell Technologies, UK) following manufacture's protocol. Then seeded at1.5 × 105 cells/ml, and cultured for 4 days in the presence of CpG oligodeoxynucleotide 2006 (10 μg/ml; Invitrogen), soluble human his-rCD40L (50ng/ml; R&D Systems, UK) and anti-poly-his (5μ g/ml; R&D Systems, UK), IL-2 (20 U/ml; R&D Systems, UK), IL-10 (50 ng/ml; Miltenyi Biotec, UK) and IL-15 (10 ng/ml; Miltenyi Biotec, UK). At day 4 of culture, the cells were washed and cultured at 2.5 × 105 cells/ml with IL-2 (20 U/ml, Milteny Biotec, UK), IL-10 (50 ng/ml), IL-15 (10 ng/m, PreproTech, UK) and IL-6 (50ng/ml; Miltenyi Biotec, UK). At day 7 of culture, cells were washed and seeded with IL-6 (50 ng/ml, PreproTech, UK), IL-15 (10 ng/ml, PreproTech, UK) and IFN-α (500 U/ml; R&D Systems) for 3 days.
Soluble human his rcd40l
Manufactured by R&D Systems
Sourced in United Kingdom
Soluble human his-rCD40L is a recombinant protein that contains the extracellular domain of human CD40 ligand fused to a histidine tag. CD40 ligand is a type II transmembrane protein that belongs to the TNF superfamily and plays a key role in immune cell activation and regulation.
Automatically generated - may contain errors
Lab products found in correlation
Cd27 cells, by STEMCELL (2 mentions)
Anti poly his, by R&D Systems (2 mentions)
Cpg oligodeoxynucleotide 2006, by Thermo Fisher Scientific (2 mentions)
Il 10, by Miltenyi Biotec (2 mentions)
Il 10, by Thermo Fisher Scientific (2 mentions)
Il 15, by Thermo Fisher Scientific (2 mentions)
Interleukin 6 (il 6), by Miltenyi Biotec (2 mentions)
Interleukin 6 (il 6), by Thermo Fisher Scientific (2 mentions)
Ifn α, by R&D Systems (2 mentions)
Interleukin 2 (il 2), by Miltenyi Biotec (2 mentions)
Il 15, by Miltenyi Biotec (2 mentions)
Interleukin 2 (il 2), by R&D Systems (2 mentions)
2 protocols using soluble human his rcd40l
1
Differentiation of Memory B Cells into Antibody-Secreting Cells
2
In Vitro Differentiation of Memory B Cells
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Memory B cells were differentiated in vitro into plasmablasts and plasma cells, following the protocol described in Jourdan et al. (24 (link)). Briefly, CD27+ Memory B cells were first purified as bulk B cells by negative selection from PBMCs by magnetic isolation followed by positive selection for CD27+ cells (Stemcell Technologies) following the manufacturer’s protocol. The cells were then seeded at 1.5 × 105 cells/ml and cultured for 4 d in the presence of CpG oligodeoxynucleotide 2006 (10 μg/ml; Invitrogen, Paisley, U.K.), soluble human his-rCD40L (50 ng/ml; R&D Systems, Abingdon, U.K.) and anti–poly-his (5 μg/ml; R&D Systems), IL-2 (20 U/ml; R&D Systems), IL-10 (50 ng/ml; Miltenyi Biotec, Surrey, U.K.), and IL-15 (10 ng/ml; Miltenyi Biotec). At day 4 of culture, the cells were washed and cultured at 2.5 × 105 cells/ml with IL-2 (20 U/ml; Miltenyi Biotec), IL-10 (50 ng/ml), IL-15 (10 ng/ml; PeproTech, London, U.K.), and IL-6 (50 ng/ml; Miltenyi Biotec). At day 7 of culture, cells were washed and seeded with IL-6 (50 ng/ml; PeproTech), IL-15 (10 ng/ml; PeproTech), and IFN-α (500 U/ml; R&D Systems) for 3 d.
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