Human Interleukin-2 Recombinant

Human Interleukin-2 Recombinant

$70.00$720.00


accession P60568


Source Optimized DNA sequence encoding Human Interleukin-2 mature chain was expressed in Escherichia Coli.
Molecular weight Native Human Interleukin-2 is generated by the proteolytic removal of the signal peptide and propeptide, the molecule has a calculated molecular mass of approximately 16kDa. Recombinant Interleukin-2 is a monomer protein consisting of 134 amino acid residue subunits, and migrates as an approximately 16kDa protein under non-reducing conditions and reducing conditions in SDS-PAGE.
Purity >95%, as determined by SDS-PAGE and HPLC
Biological Activity The ED(50) was determined by the dose-dependent stimulation of the proliferation of monkeyMBr-5 cells was found to be in the range of.0-40.0 ng/ml.

Protein Sequence MYRMQLLSCI ALSLALVTNS APTSSSTKKT QLQLEHLLLD LQMILNGINN YKNPKLTRML TFKFYMPKKA TELKHLQCLE EELKPLEEVL NLAQSKNFHL RPRDLISNIN VIVLELKGSE TTFMCEYADE TATIVEFLNR WITFCQSIIS TLT
Endotoxin Endotoxin content was assayed using a LAL gel clot method. Endotoxin level was found to be less than.1 ng/µg(1EU/µg).
Presentation Recombinant Human Interleukin-2 was lyophilized from a.2 μm filtered PBS solution.
Reconstitution A quick spin of the vial followed by reconstitution in distilled water to a concentration not less than.1 mg/mL. This solution can then be diluted into other buffers.
Storage The lyophilized protein is stable for at least years from date of receipt at -20° C. Upon reconstitution, this cytokine can be stored in working aliquots at° -° C for one month, or at -20° C for six months, with a carrier protein without detectable loss of activity. Avoid repeated freeze/thaw cycles.
Usage This cytokine product is for research purposes only.It may not be used for therapeutics or diagnostic purposes.


Interactor P14784 IL2RB_HUMAN
Interactor P31785 IL2RG_HUMAN
Biological Process Immunity
Molecular function Cytokine
Molecular function Growth-factor

Methods

Phenotype of clinical grade LAKs and DCs.

IL-2
  • Phenotype analysis was performed on CD14− PBMCs before and after culture for 5 days in IL-2 to generate LAKs.

Derivation of γδ T Cells

  • Human peripheral blood was collected (30 ml) from adult healthy donors after obtaining the IRB approval from the Ohio State University Medical Center and obtaining written consents from donors.
  • The ethic committee has also approved the procedure and records are saved in the laboratory logbook.
  • Freshly collected blood was processed to isolate peripheral blood mononuclear cells (PBMC) following the similar protocol published earlier

2.5. Transduction of PBMCs and T Cells

  • Peripheral blood monocytes (PBMCs) were from an HLA-A2, healthy human.
  • T cells were obtained from anti-CD3 conjugated magnetic beads .
  • The PBMCs and T cells were cultured in AIM-V and interleukin-2 (IL-2; PeproTech, , , ) at 300 IU/mL.
  • For medium'>OKT3 stimulation, the cells were placed initially in either a medium with anti-CD3 antibody, medium'>OKT3 at 50 ng/mL or in an medium'>OKT3 medium after transduction at the initial changing of the culture medium in the presence of IL-2.
  • For transduction of the PBMCs or T cells, 1 × 106 cells were adjusted to a final volume of 1 mL in a 24-well, tissue culture-treated plate with the viral supernatant and(8 mg/mL, , , ).
  • The cells were transduced by centrifugation of the plates at 1000 g for 1.5 hours at 32°C.
  • The plates were placed in a 37°C,…
  • Peripheral blood monocytes (PBMCs) were from an HLA-A2, healthy human.
  • T cells were obtained from anti-CD3 conjugated magnetic beads .
  • The PBMCs and T cells were cultured in AIM-V and interleukin-2 (IL-2; PeproTech, , , ) at 300 IU/mL.
  • For medium'>OKT3 stimulation, the cells were placed initially in either a medium with anti-CD3 antibody, medium'>OKT3 at 50 ng/mL or in an medium'>OKT3 medium after transduction at the initial changing of the culture medium in the presence of IL-2.
  • For transduction of the PBMCs or T cells, 1 × 106 cells were adjusted to a final volume of 1 mL in a 24-well, tissue culture-treated plate with the viral supernatant and(8 mg/mL, , , ).
  • The cells were transduced by centrifugation of the plates at 1000 g for 1.5 hours at 32°C.
  • The plates were placed in a 37°C, humidified, 5% CO2 incubator overnight, and the medium was replaced the next day.

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IL-7 induces partial reactivation of latent HIV-1 in cultured TCM.

IL-2
  • Mock or DHIV latently infected, cultured TCM were incubated in the presence of IL-2, IL-7 or a combination of IL-2 and IL-7 (IL-2+IL-7) or costimulated with antibodies to CD3 and CD28 (αCD3/αCD28) and assessed for intracellular p24Gag by flow cytometry (1st Reactivation).

Cell isolation and culture

  • Peripheral blood mononuclear cells (PBMC) from CLL patients or healthy donors were separated from heparinized venous blood by Ficoll density gradient centrifugation.
  • The PBMC used in this study were generally greater than 90% CLL B cells as determined by CD19/CD5 positivity.
  • Leukemic cells were cultured in serum-free adoptive immunotherapy media-V medium'>(AIM-V) medium .
  • For CLL B cell activation experiments, cells were cultured for 5 days in AIM-V media or AIM-V media supplemented with 2.5 µg/ml CpG ODN 2006 (provided by core Mayo Clinic facility) with or without 94 IU/ml human IL-2 , 10 ng/ml human IL-15 , and 13 µM beta-mercaptoethanol.
  • Further CLL B cell activation experiments utilized 12-O-tetradecanoylphorbol-13-acetate (TPA) at 10 ng/ml or interferon-α 2b at 1000 IU/ml.
  • Human C4+T cells were isolated from healthy donor PBMC by immunomagnetic selection using T cell enrichment kits and incubated with CLL B cells
  • Peripheral blood mononuclear cells (PBMC) from CLL patients or healthy donors were separated from heparinized venous blood by Ficoll density gradient centrifugation.
  • The PBMC used in this study were generally greater than 90% CLL B cells as determined by CD19/CD5 positivity.
  • Leukemic cells were cultured in serum-free adoptive immunotherapy media-V medium'>(AIM-V) medium .
  • For CLL B cell activation experiments, cells were cultured for 5 days in AIM-V media or AIM-V media supplemented with 2.5 µg/ml CpG ODN 2006 (provided by core Mayo Clinic facility) with or without 94 IU/ml human IL-2 , 10 ng/ml human IL-15 , and 13 µM beta-mercaptoethanol.
  • Further CLL B cell activation experiments utilized 12-O-tetradecanoylphorbol-13-acetate (TPA) at 10 ng/ml or interferon-α 2b at 1000 IU/ml.
  • Human C4+T cells were isolated from healthy donor PBMC by immunomagnetic selection using T cell enrichment kits and incubated with CLL B cells at a ratio of 1∶2 in plates pre-coated with PBS containing 10 µg/ml of anti-C3 and anti-C28 (B) stimulating antibodies.
  • All cells were maintained in appropriate media at 37°C in an atmosphere containing 95% air and 5% CO2.

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2.4. Coculture of CHO/IDO Cells and CD3+ T Cells

  • The CD3+ T cells in peripheral blood mononuclear cells (PBMCs) of breast cancer patients were purified using Human Pan T-cell Isolation Kit II according to the manufacturer's instructions.
  • 1 × 105 CHO/IDO cells and CHO/EGFP cells were seeded in a 24-well plate and cocultured with 2 × 106 purified T cells in complete RPMI 1640 medium supplemented with 10% FBS and 50 U/mL rhIL-2 at 37°C in a 5% CO2 incubator.
  • Unstimulated T cells cocultured in complete RPMI 1640 medium supplemented with 10% FBS and 50 U/mL rhIL-2 were used as control.
  • The nonadherent T cells under different treatments were harvested 7 days later for flow cytometry analysis, quantitativee real time RT-PCR, and Western Blot analysis.

Stimulation of antiviral T cells with HSP70, CMVpp65495-503 peptide, and HSP70/CMV-PC

  • T-cell stimulation was performed using samples from 16 A2/CMV-pentamer-positive donors.
  • Peripheral blood mononuclear cells (PBMCs) were isolated by discontinuous gradient centrifugation, washed twice in sterile PBS, resuspended at a concentration of 1 × 107 cells/ml in medium'>RPMI1640 culture medium , and then supplemented with 10% heat-inactivated human AB serum (C.C.pro, Neustadt, ) and 100 U/ml IL-2 .
  • Cells were stimulated in a 24-well plate with either 10 μg/ml HSP70, 10 μg/ml CMVpp65495-503 peptide, or 10 μg/ml HSP70/CMV-PC.
  • HSP70-peptide-binding buffer was used as negative control.

Retroviral delivery of MART-1 specific TcRs.

100 U IL-2
  • IL-2 release was monitored by ELISA assay, and plotted as pg of IL-2 per mL of medium.

Cytokine profile of tonsil MNCs following S. pneumoniae antigen stimulation.

25 U/ml IL-2
  • Cytokine (IL-10, IFNγ, TNFα, IL-2 and IL-17) production by tonsil MNCs following stimulation with Ply (grey bars, SEM are shown by error bars)) was assessed by quantifying cytokine levels in cell culture supernatants (n = 8) at 7 days post cell stimulation by Luminex assay.

Ex vivo culture of human peripheral blood T cells

  • For regulatory T cell culture, human enriched T cells were co-cultured with MSCs or SB623 cells in the presence of human interleukin-2 (IL-2) at a 10:1 T cells to MSC or SB623 cell ratio for 7 days followed by cell surface staining for CD4, a helper T cell marker and CD25, the IL-2 receptor alpha chain.
  • For FoxP3 intracellular staining, cells were fixed and permeabilized with CytoFix/Perm (eBioscience).
  • PE-conjugated antibody against FoxP3 (clone PCH101, eBioscience) was used at 1:50 dilution and flow cytometry analysis was done gating on lymphocytes.
  • For assessment of constitutive IL-10 production, intracellular staining with fluorochrome conjugated antibody against IL-10 was performed without PMA/Io stimulation on7.