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Ficoll

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Ficoll is a high-molecular-weight, synthetic polymer used as a density gradient medium for the separation and purification of cells, subcellular organelles, and macromolecules. It is commonly used in the isolation of mononuclear cells from whole blood samples.

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Lab products found in correlation

Lympholyte, by Cedarlane (4 mentions) Adam mc, by NanoEnTek (3 mentions) L glutamine, by Euroclone (3 mentions) α minimal essential medium α mem, by Lonza (2 mentions) Recombinant human macrophage colony stimulating factor rh mcsf, by Thermo Fisher Scientific (2 mentions) Fetal bovine serum (fbs), by Euroclone (2 mentions) Rpmi 1640, by Thermo Fisher Scientific (2 mentions) Fetal bovine serum (fbs), by Biosera (2 mentions) Rpmi 1640 medium, by Euroclone (2 mentions) Fetal bovine serum (fbs), by Thermo Fisher Scientific (2 mentions) Luzindole, by Merck Group (1 mentions) Tamoxifen tmx, by Merck Group (1 mentions) Resveratrol, by ChromaDex (1 mentions) 2 7 dichlorofluorescin diacetate dcfh da, by Merck Group (1 mentions) Dulbecco s phosphate buffered saline dpbs, by Thermo Fisher Scientific (1 mentions) Quercetin, by Merck Group (1 mentions) Gentamicin, by Merck Group (1 mentions) Phorbol 12 myristate 13 acetate pma, by Merck Group (1 mentions) Ascorbic acid, by Tokyo Chemical Industry (1 mentions) Edta tube, by BD (1 mentions)

25 protocols using ficoll

1

PBMC Isolation and T Cell Analysis

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Whole blood from 50 Italian Healthy Controls (HCs) was collected by venipuncture in Vacutainer tubes containing EDTA (Ethylene diamine tetracetic acid) (BD Vacutainer, San Diego, CA). Peripheral Blood Mononuclear Cells (PBMCs) were isolated by density gradient centrifugation on Ficoll (Cedarlane Laboratories Limited, Hornby, Ontario, Canada) and counted with the automated cell counter ADAM-MC (Digital Bio, NanoEnTek Inc., Korea), which allows for discrimination of viable from non-viable cells. PBMCs isolated from 8 subjects (4 HomoA and 4 HomoB) were incubated for 24-h with/without recombinant human (rh) ERAP2, in order to analyse T cell apoptosis, proliferation, activation, and maturation.
The Ethical Committee of the Don C. Gnocchi Foundation IRCCS approved the study (Prot. N°10/2018/CE_FdG/SA). All the donors signed an informed consent form, in accordance with the Declaration of Helsinki.
Saulle I., Ibba S.V., Torretta E., Vittori C., Fenizia C., Piancone F., Minisci D., Lori E.M., Trabattoni D., Gelfi C., Clerici M, & Biasin M. (2019). Endoplasmic Reticulum Associated Aminopeptidase 2 (ERAP2) Is Released in the Secretome of Activated MDMs and Reduces in vitro HIV-1 Infection. Frontiers in Immunology, 10, 1648.
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2

Hypoxic Isolation of Murine Immune Cells

Cited 2 times
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All BM and PB collections were performed in a hypoxia chamber, maintained at 3% O2, 5% CO2, and N2 balance. All reagents and supplies were equilibrated in the 3%O2 hypoxia chamber for at least 16 hours prior to cell collection time (15 (link)). To isolate PB mononuclear cells (PBMCs), PB was collected by intracardiac aspiration with heparinized syringes immediately after euthanizing mice. PBMC were isolated using lymphocyte mammal Ficoll (CEDARLANE, Burlington, ON, Canada) with gradient centrifugation isolation. All tubes were sealed with parafilm to maintain the 3% O2 during centrifugation outside the hypoxia chamber. BM was harvested by flushing femurs with PBS. All subsequent techniques, flow cytometry staining, and fixation, HPC colony assays, and injections for transplantation were performed in the hypoxia chamber. For ambient air control groups, we removed half of the cells from the hypoxia chamber to ambient air for at least 60 minutes before applying equivalent techniques in ambient air. We had previously shown that cells collected immediately in air showed similar effects to cells collected in hypoxia and then equilibrated to air for 1 hour (15 (link)). Cells assessed by colony-forming cell assays were incubated at 5% O2, 5% CO2 regardless of how cells were collected and processed, as this allows detection of optimal growth (16 (link)).
Aljoufi A., Cooper S, & Broxmeyer H.E. (2020). Collection and Processing of Mobilized Mouse Peripheral Blood at Lowered Oxygen Tension Yields Enhanced Numbers of Hematopoietic Stem Cells. Stem cell reviews and reports, 16(5), 946-953.
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3

PBMC Isolation from Healthy Donors

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Venous blood samples were obtained from eight healthy male nonsmoker donors aged between 25 and 35 years who were included in two different experiments. Afterward, peripheral blood mononuclear cells (PBMCs) were isolated from diluted blood using Ficoll (Cedarlane, Canada) density gradient centrifugation. The separated layer was then washed twice with phosphate-bufferedsaline. Collection of the blood samples was approved by the ethics committee of Mashhad University of Medical Sciences (approval code: IR.MUMS.MEDICAL.REC.1397.654) and written informed consent was obtained from all donors before blood collection.
Mollaee P.F., Azimian H., Ghadim N.Z., Dolat E., Sheykhoo A, & Bahreyni-Toossi M.T. (2023). The role of intrinsic radiosensitivity in the low-dose adaptive response induction in human peripheral blood mononuclear cells. Journal of cancer research and therapeutics, 19(Suppl 2).
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4

Melatonin and Calmodulin Receptor Antagonists

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Luzindole (antagonist for melatonin MEL-a receptor, 5 mg/single), Sigma (USA).

Tamoxifen (TMX) (receptor antagonist of calmodulin, 1 g/single), Sigma (USA).

Ficoll to separate and purify the blood platelets, Cedarlane, Ltd., number: CL5020-6.

CaM Rat Kit to measure the level of calmodulin in platelets of rats from each group, Wuhan Gene Biological Technology Co., Ltd.

Absolute alcohol, Hefei Sanding Chemical Co., Ltd.

Wu J.Z., Wu W.H., He L.J., Ke Q.F., Huang L., Dai Z.S, & Chen Y. (2016). Effect of Melatonin and Calmodulin in an Idiopathic Scoliosis Model. BioMed Research International, 2016, 8460291.
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5

Isolation and Differentiation of Human Macrophages

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Macrophages were obtained from peripheral blood mononuclear cells (PBMCs). PBMCs were isolated by means of density gradient centrifugation (Ficoll 1.077 g/mL; Lympholyte, Cedarlane Laboratories Ltd., Uden, The Netherlands); diluted at 1 × 106 cells/mL in α-Minimal Essential Medium (α-MEM) (Lonza, Verviers, Belgium) supplemented with 10% fetal bovine serum (FBS) (Euroclone, Siziano, Italy), 100 IU/mL penicillin, and 100 g/mL streptomycin and L-glutamine (Gibco Limited, Uxbridge, UK); and plated in 24-well Cell Culture Multiwell. To obtain fully differentiated human macrophages, the PBMCs were cultured for 15 days (about 2 weeks) in the presence of 25 ng/mL recombinant human macrophage colony-stimulating factor (rh-MCSF) (Peprotech, London, UK). The culture medium was replaced twice a week. Cells were cultured at 37 °C in a humidified atmosphere with 5% CO2. Macrophages were treated with Res, Cur, and OE-Lyc for 24 h after complete differentiation. Cells were then harvested for protein extraction, and cell culture supernatants were collected to perform iron assays and ELISA.
Di Paola A., Marrapodi M.M., Pota E., Colucci Cante R., Rana D., Giliberti G., Di Feo G., Ahmed S., Roberti D., Nigro R., Rossi F, & Argenziano M. (2024). Role of Nutraceuticals in Counteracting Inflammation in In Vitro Macrophages Obtained from Childhood Cancer Survivors. Cancers, 16(4), 714.
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6

Ionizing Radiation Effects on PBMCs

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Peripheral blood mononuclear cells (PBMCs) were separated from the 5 ml EDTA blood samples by density-gradient centrifugation using Ficoll (Cedar lane Lab, Canada) according to the manufacturer’s instructions. PBMCs were washed twice with physiological phosphate buffer saline (PBS) and finally re-suspended in the 10 ml culture medium containing RPMI 1640 (GIBCO, Germany), 10% fetal bovine serum (FBS, Biosera, France), 100 Iu/ml penicillin, and 0.1 µg/ml streptomycin. Each sample was divided into three flasks including 1 and 2 Gy radiation and control group. Lymphocytes were irradiated by 6 MV X-rays to deliver 1 and 2 Gy in-vitro on ice.
Azimian H., Dayyani M., Toossi M.T, & Mahmoudi M. (2018). Bax/Bcl-2 expression ratio in prediction of response to breast cancer radiotherapy. Iranian Journal of Basic Medical Sciences, 21(3), 325-332.
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7

Antioxidant Potential Evaluation of Novel Compounds

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The novel compounds 1–9 were purchased from Vitas-M, 10 and 11 from Key Organics, 12–14 from ChemDiv, 15 from MolMall, 16 from MolPort, 17 from ChemBridge, and 18–21 from Sigma Aldrich, while the standard antioxidants aesculin, ascorbic acid, curcumin, scopoletin, Trolox and gallic acid were purchased from TCI Europe, resveratrol from ChromaDex, and quercetin from Sigma Aldrich. The structures of the novel compounds and standard antioxidants are presented in Fig 4.
2,2-diphenyl-1-picrylhydrazyl (DPPH), 2-deoxyribose, β-carotene, linoleic acid, lipoxidase from Glycine max, trypan blue solution, gentamicin, phorbol 12-myristate 13-acetate (PMA) and 2',7'-dichlorofluorescin diacetate (DCFH-DA) were all purchased from Sigma-Aldrich. Sterile dimethyl sulfoxide (DMSO) was obtained from WAK-Chemie Medical GmbH, fetal bovine serum (FBS), glutamine (GlutaMAX) and Dulbecco’s Phosphate-Buffered Saline (DPBS) from Gibco, Ficoll from Cedarlane, H2O2 from Merck, and RPMI 1640 medium from PAA Laboratories GmbH. All other chemicals (EDTA, FeCl3, methanol, tween 20, chloroform, thiobarbituric acid, trichloroacetic acid, NaOH, KH2PO4, and K2HPO4) were of reagent grades and obtained from the local chemical vendors in Slovenia.
Martinčič R., Mravljak J., Švajger U., Perdih A., Anderluh M, & Novič M. (2015). In Silico Discovery of Novel Potent Antioxidants on the Basis of Pulvinic Acid and Coumarine Derivatives and Their Experimental Evaluation. PLoS ONE, 10(10), e0140602.
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8

SARS-CoV-2 Antigen Stimulation of PBMCs

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Whole blood was collected from both patients in EDTA tubes (BD Vacutainer, San Diego, CA). Samples from patient 1 were obtained at three different points (T0 = pre-transplantation, T1 = 1 day post-transplantation, and T7 = 7 days post-transplantation). PBMCs were isolated by density gradient centrifugation on Ficoll (Cedarlane Laboratories Limited, Hornby, ON, Canada) and viable cells were counted with the automated cell counter ADAM-MC (Digital Bio, NanoEnTek Inc., Seoul, KR, Korea). PBMCs were resuspended at the concentration of 1 × 106/mL in RPMI 1640 medium (Euroclone, Milan, Italy) supplemented with 10% fetal bovine serum, 1% of L-glutamine (LG), and 2% pen-streptomycin. Subsequently, PBMCs were stimulated with 500 ng/mL nucleocapsid- (N) and spike- (S) specific SARS-CoV-2 antigens (Novatein Biosciences, Cambridge, MA, USA). For gene expression and cytokine analyses, cells were harvested 10 h after stimulation and cytokine content was quantified on cell culture supernatants.
Croci G.A., Vaira V., Trabattoni D., Biasin M., Valenti L., Baselli G., Barberis M., Guerini Rocco E., Gregato G., Scandroglio M., Fominskiy E., Palleschi A., Rosso L., Nosotti M., Clerici M, & Ferrero S. (2021). Emergency Lung Transplantation after COVID-19: Immunopathological Insights on Two Affected Patients. Cells, 10(3), 611.
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9

Saliva and Plasma Biospecimen Processing

Cited 1 time
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Participants were asked not to eat, drink, or smoke for at least 30 min prior to saliva collection. Saliva was incubated at 56 °C for 10 min and centrifuged at 6000× g for 10 min. Supernatants were stored at −80 °C until use. Plasma was obtained by centrifugation of whole blood at 1200× g for 10 min and stored at −20 °C until use.
PBMCs were isolated from whole blood by density gradient centrifugation using Ficoll (Cedarlane Laboratories Limited, Hornby, ON, Canada), as previously described [10 (link)], and viable cells were counted with the cell counter ADAM-MC (Digital Bio, NanoEnTek Inc., Seoul, Republic of Korea).
Saulle I., Gidaro A., Donadoni M., Vanetti C., Mutti A., Romano M.E., Clerici M., Cogliati C, & Biasin M. (2024). Immunological Profiles in Parry–Romberg Syndrome: A Case–Control Study. Journal of Clinical Medicine, 13(5), 1219.
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10

PBMC Isolation and Culture Protocol

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Fifty milliliter whole blood samples were collected in individual tubes containing Ethylene Diamine Tetra-acetic acid (EDTA) from male volunteers. PBMCs were separated by Ficoll (Cedarlane) according to the recommendations of the manufacturer. The mononuclear cell layer was removed and washed in Phosphate Buffer Saline (PBS). The cells were counted and re-suspended in a 10 ml culture medium, including RPMI 1640 (Gibco) containing 20% FBS (Biosera), 1% of 200 mM L-glutamine (Biosera), 100 Iu/ml penicillin, 0.1 μg/ml Streptomycin. Cells were transferred to four 25 cm3 flasks (Spl) in a final cell concentration of 1 × 106 cells/ml.
Azimian H., Bahreyni-Toossi M.T., Rezaei A.R., Rafatpanah H., Hamzehloei T, & Fardid R. (2015). Up-regulation of Bcl-2 expression in cultured human lymphocytes after exposure to low doses of gamma radiation. Journal of Medical Physics / Association of Medical Physicists of India, 40(1), 38-44.
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