Cells were stimulated with human recombinant IL4 (BD Pharmingen), IL13 (Peprotech) or medium for 4, 8, and 24 hours. STAT6 expression and phosphorylation were determined by Western blot (anti-pSTAT6, BD Biosciences; anti-STAT6 Santa Cruz Biotechnology).
Human recombinant il 4
Manufactured by BD
Sourced in United States
Human recombinant IL-4 is a cytokine produced using recombinant DNA technology. It is a protein that plays a role in the regulation of immune responses.
Automatically generated - may contain errors
Lab products found in correlation
Annexin 5, by BD (2 mentions)
Interleukin 4 (il 4), by BD (2 mentions)
Fetal calf serum (fcs), by Cambrex (2 mentions)
Propidium iodide, by BD (2 mentions)
Anti pstat6, by BD (1 mentions)
Anti stat6, by Santa Cruz Biotechnology (1 mentions)
Il 13, by Thermo Fisher Scientific (1 mentions)
Bio whittaker, by Cambrex (1 mentions)
Rpmi 1640, by Cambrex (1 mentions)
5 protocols using human recombinant il 4
1
IL4/IL13 Induced STAT6 Activation
2
Apoptosis Induction in B-Cell Leukemia
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Following purification, three fractions of the purified CLL and NBC were processed: a) at time zero (“Pre”); b) after being cultured for 18 hours in RPMI-1640 medium supplemented with 10% fetal calf serum (Cambrex, East Rutherford, NJ) (“Ctrl”); and c) as b, but with adding 10 ng/mL of human recombinant IL-4 (BD Biosciences, San Diego, CA) (“IL-4”). In the absence of sufficient material, the Ctrl culture was not carried out in 3 NBC (NBC06, NBC07, and NBC08). In selected patients, additional fractions were treated with IL-4 plus InSolution NF-kB activation inhibitor [6-Amino-4-(4-phenoxyphenylethylamino)quinazoline] (Merck, Nottingham, UK) at 1 µM and 10 µM. Apoptosis of the cultured cells was determined by dual labelling with annexin V and propidium iodide (BD Biosciences), and flow cytometry analysis.
3
Isolation and Culture of CLL B Cells
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Whole blood samples from 5 volunteer donors and 12 chronic lymphocytic leukemia (CLL) patients, and spleen samples from 5 cadaveric transplant donors, were previously used [12] (link). The study was approved by the Institutional Review Board of the Hospital Virgen de la Arrixaca and written informed consent was obtained according to the Helsinki Declaration guidelines. CLL B cells were cultured in RPMI-1640 medium with 10% fetal calf serum (Biowhittaker, Cambrex, East Rutherford, NJ), 50 U/mL penicillin, 50 U/mL streptomycin, 2,5 μg/mL amphotericin B, and 2 mM L -glutamine (‘RPMI-C’, where ‘C’ means ‘complete’) and 10 ng/mL human recombinant IL-4 (BD Pharmingen, BD Biosciences, San Diego, CA).
4
B-cell Neoplasm Isolation and IL-4 Stimulation
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Whole blood from seven patients with different B-cell neoplasms was obtained including two patients with mantle cell lymphoma (MCL) and one patient from each of these entities: prolymphocytic leukemia (PLL), diffuse large B-cell lymphoma (DLBCL), non-Hodgkin lymphoma (NHL), plasma cell leukemia (PCL), and CLL, diagnosed according to the guidelines of the World Health Organization (WHO) [20 (link)]. All the patients had leukocytosis, and none were receiving chemotherapy at the time of sampling. The samples were processed for isolation of B cells using RossetteSep Enrichment Cocktails (StemCell Technologies, Vancouver, Canada). Cell aliquots were cultured for 24 h without or with 10 ng/mL human recombinant IL-4 (BD Pharmingen, BD Biosciences, San Diego, CA, USA) in RPMI-1640 medium with the same supplements described in Section 2.1 .
5
Apoptosis Profiling of CLL and NBC Cells
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Following purification, three fractions of the purified CLL and NBC were processed: a) at time zero (“Pre”); b) after being cultured for 18 hours in RPMI-1640 medium supplemented with 10% fetal calf serum (Cambrex, East Rutherford, NJ) (“Ctrl”); and c) as b, but with adding 10 ng/mL of human recombinant IL-4 (BD Biosciences, San Diego, CA) (“IL-4”). Apoptosis of the cultured cells was determined by dual labelling with annexin V and propidium iodide (BD Biosciences), and flow cytometry analysis.
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