Transferrin

Manufactured by Merck Group

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Transferrin is a laboratory product used as a critical component in cell culture media. It is a glycoprotein that plays a key role in the transport and delivery of iron to cells. Transferrin functions to bind and carry iron ions, facilitating their uptake and utilization by cells in vitro.

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Lab products found in correlation

Close spelling variants of this product

Human transferrin (116 citations) Human holo-transferrin (59 citations) Insulin-transferrin-sodium selenite (ITS, (13 citations) Insulin-transferrin-selenium premix (10 citations) Insulin-transferrin-selenium supplement (9 citations) Insulin-transferrin-selenium solution (6 citations) Insulin-Transferrin-Selenium supplement (ITS; (5 citations) Bovine apo-transferrin (4 citations) Insulin-transferrin-sodium selenite media (3 citations) Iron-saturated transferrin (3 citations)

602 protocols using transferrin

Cultivation of Human Colonic and Macrophage Cells

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Human colonic epithelial cells (HCEC‐1CT, CkHT‐039‐0229; Evercyte GmbH, Vienna) were routinely grown at 37°C under a humidified atmosphere of 7% CO₂ in DMEM:199 (4:1/Biochrome, Germany) supplemented with 2% cosmic calf serum (HyClone, Logan, UT), EGF (25 ng/ml), hydrocortisone (1 g/ml), insulin (10 g/ml), transferrin (2 g/ml), and sodium selenite (5 nM; all from Sigma‐Aldrich, St. Louis, MO). Differentiation of cells toward colonic epithelial cells (described by Roig et al., 2010) was induced by culturing cells for 48 hr in DMEM:199 (4:1) supplemented with 0.1% cosmic calf serum, EGF (1.25 ng/ml), hydrocortisone (1 µg/ml), insulin (10 µg/ml), transferrin (2 µg/ml), sodium selenite (5 nM), and GSK‐2 inhibitor IX (5 µm; Merck).
U937 cells (ATCC CRL 1593) were cultivated in Roswell Park Memorial Institute 1640 media (Biochrom, Germany) containing 10% heat‐inactivated fetal calf serum and 4 mM l‐glutamine (Sigma‐Aldrich). Differentiation of cells toward macrophage‐like cells was induced by culturing 7 × 10⁵ cells/ml in medium containing 100 nM phorbol 12‐myristate 13‐acetate for 24 hr. The medium was changed to routine medium and cells were cultivated for further 48 hr until the cells attached to the surface showing the development of a dendritic‐like morphology.

Schwestka J., Tschofen M., Vogt S., Marcel S., Grillari J., Raith M., Swoboda I, & Stoger E. (2020). Plant‐derived protein bodies as delivery vehicles for recombinant proteins into mammalian cells. Biotechnology and Bioengineering, 117(4), 1037-1047.

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Immortalized motoneuronal cell line NSC34 transfection

Cited 2 times

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The immortalized motoneuronal cell line NSC34 (58 (link),59 (link)) is routinely used in our laboratory (24 (link),37 (link),39 (link)–41 (link),52 (link),54 (link),60 (link)–62 (link)) and has been transfected with Lipofectamine (Life Technologies Corporation, Carlsbad, CA, USA)/transferrin (Sigma-Aldrich), as previously described (39 (link),40 (link),60 (link)), using 0.6 µg of plasmid DNA, 4 µl of transferrin solution and 2 µl of Lipofectamine.
Rat adrenal pheochromocytoma (PC12) cells stable transfected with the plasmid encoding AR.Qn (n010, 112) express AR under the control of a Tet-On promoter responsive to 1 µg/ml of doxycycline [produced (and kindly provided) by Professor D. Merry (TJU, Philadelphia, PA, USA)]. In the experiments involving steroid hormone treatments, the fetal bovine serum (FBS) was replaced with charcoal-stripped FBS and the horse serum (HS) with charcoal-stripped HS, to eliminate endogenous steroids (24 (link),40 (link),63 (link)).

Giorgetti E., Rusmini P., Crippa V., Cristofani R., Boncoraglio A., Cicardi M.E., Galbiati M, & Poletti A. (2014). Synergic prodegradative activity of Bicalutamide and trehalose on the mutant androgen receptor responsible for spinal and bulbar muscular atrophy. Human Molecular Genetics, 24(1), 64-75.

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Erythroblast differentiation regulation

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GFP⁺ lentivirus-transduced erythroblasts (CD71⁺/Ter119^-) were seeded at a density of 1 × 10⁶ cells/mL in StemPro-34 SFM (ThermoFisher) supplemented with l-glutamine (1%; ThermoFisher), Epo (Peprotech; 10 U/mL) and transferrin (Sigma-Aldrich; 1 mg/mL) as previously published^{36 (link)}. The GFP⁺ cells were cultured for 48 h in the presence and absence of ICG001 (Tocris) and JW74 (Tocris). Half media exchange was performed after 24 h, and after 48 h the cells were collected, stained with CD71 and Ter119 antibodies and analyzed using flow cytometry.
For experiments involving FL cells, FACS sorted CD71⁺Ter119^- FL cells were seeded at a density of 0.5 × 10⁶ cells/mL StemPro-34 SFM (ThermoFisher) supplemented with l-glutamine (1%; ThermoFisher), Epo (Peprotech; 10 U/mL) and transferrin (Sigma-Aldrich; 1 mg/mL). The FL cells were cultured for 48 h in the presence and absence of ICG001 (Tocris) and JW74 (Tocris). Half media exchange was performed after 24 h, and after 48 h the cells were collected, stained with CD71 and Ter119 antibodies and analyzed using flow cytometry.

Rothberg J.L., Maganti H.B., Jrade H., Porter C.J., Palidwor G.A., Cafariello C., Battaion H.L., Khan S.T., Perkins T.J., Paulson R.F., Ito C.Y, & Stanford W.L. (2018). Mtf2-PRC2 control of canonical Wnt signaling is required for definitive erythropoiesis. Cell Discovery, 4, 21.

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Inducible NSC34 Motor Neurone Model

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Immortalized mouse motor neurone cell line NSC34 (54 (link),55 ) is routinely cultured in our laboratory (45 (link),46 (link),56–65 (link)). Cells were plated at 80,000 cells/well in 12-well multiwell plates for Western Blot (WB) and Filter Retardation Assay (FRA) analysis; for immunohistochemistry analysis, cells were plated at 35,000 cells/well in 24-well multiwell plate with coverslips. 24 h after plating, the cells were transiently transfected with lipofectamine (Life Technologies Corporation)/transferrin (Sigma-Aldrich), following the manufacture’s instructions (1 μg of plasmid DNA, 4 μl of transferrin solution and 2 μl of lipofectamine per well of 12-well multiwell plate). Inducible stably-transfected GFP-TDPs-NSC34 cell lines were obtained by transiently transfecting a TR4 NSC34 cell line (66 (link),67 (link)), kindly provided by Dr. E. Garattini (Mario Negri Institute, Milan, Italy), with pcDNA5/TO-GFP-TDPs plasmids and selecting positive clones with 200 μg/ml hygromycin for 3 weeks. GFP-TDPs-NSC34 selected clones were cultured in low-glucose DMEM (Euroclone, Pero, Milan, Italy) supplemented with 5% of tetracycline-free serum (Euroclone), 100 μg/ml hygromycin (Euroclone) as described in Locatelli et al. 2012 (66 (link)). For FRA analyses, cells were plated at 70,000 cells/well in 12-well multiwell plate and induced with 1 μg/ml doxycycline for 72 h.

Crippa V., Cicardi M.E., Ramesh N., Seguin S.J., Ganassi M., Bigi I., Diacci C., Zelotti E., Baratashvili M., Gregory J.M., Dobson C.M., Cereda C., Pandey U.B., Poletti A, & Carra S. (2016). The chaperone HSPB8 reduces the accumulation of truncated TDP-43 species in cells and protects against TDP-43-mediated toxicity. Human Molecular Genetics, 25(18), 3908-3924.

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Protein-Iron Complex Characterization

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Standard buffer consisted of 25 mM sodium bicarbonate (Fisher), 20 mM Tris-base (Fisher), and 10 mM NaCl (EMD), prepared in high purity water and adjusted to pH 7.4 using trace-grade HCl (Fisher). Stock solutions of transferrin (Athens Research and Technology) (76 μM) and bovine serum albumin (Sigma) (1.2 mM) were prepared in the standard buffer. A stock solution of Fe^III citrate (70 μM) was prepared by mixing 40 mM ferrous ammonium sulfate (Fisher) with 120 mM sodium citrate (Fisher), and adjusting the pH to 5 using citric acid (Acros Organics) (final concentrations obtained using deionized water). Iron solutions were mixed with select protein samples, transferrin:iron at molar ratio 1:1 and albumin:iron at molar ratio 0.058:1. Protein and iron solutions were mixed, incubated for 1 hr, then passed through a 0.2 μm filter (VWR). transferrin ± Fe and albumin ± Fe solutions were mixed 1:1 with each other or diluted 1:1 with standard buffer. A solution of alcohol dehydrogenase (Sigma) was prepared (10 mg/mL) in standard buffer, and mixed 1:1:1 with Fe^III transferrin (prepare 1:1 Fe^III citrate) and albumin solutions (in this sample, only transferrin had Fe^III citrate). Ferritin (Sigma; 2 mg/mL) was prepared in standard buffer except lacking bicarbonate. These solutions were then passed though the Superdex 200 GL column as described above.

Dziuba N., Hardy J, & Lindahl P.A. (2019). Low-molecular-mass iron complexes in blood plasma of iron-deficient pigs do not originate directly from nutrient iron. Metallomics : integrated biometal science, 11(11), 1900-1911.

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Culturing Murine Cortical Neurons

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Cortical neurons were isolated from P1–2 wild-type mice and plated at 50,000–75,000 cells per coverslip on poly-ornithine-coated coverslips (12 mm). Neurons were plated in media containing DMEM supplemented with 5 g/L glucose, 100 mg/L Transferrin (Millipore), 10% FBS, 2% B-27 (Thermo Fisher), 1% Glutamax (Thermo Fisher) and 0.25 g/L insulin. At DIV 3–4, 50% of the plating media was removed and exchanged for feeding media containing media supplemented with 4 μM cytosine β-d-arabinofuranoside (Ara-C).

Acosta-Ruiz A., Gutzeit V.A., Skelly M.J., Meadows S., Lee J., Parekh P., Orr A.G., Liston C., Pleil K.E., Broichhagen J, & Levitz J. (2019). Branched photoswitchable tethered ligands enable ultra-efficient optical control and detection of G protein-coupled receptors in vivo. Neuron, 105(3), 446-463.e13.

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Myelination of iPSC-derived Nociceptors

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Schwann cell co-cultures were prepared as previously described (Clark et al., 2017 (link)). In brief, 30,000 rat Schwann cells were added to 30 DIV iPSC-derived nociceptors in Schwann cell basal medium [DMEM/F12 (ThermoFisher), 5 mg/ml insulin (Sigma), 100 mg/ml transferrin (Millipore), 25 ng/ml recombinant-human NGF (Peprotech), 25 ng/ml Selenium (Sigma), 25 ng/ml thyroxine (Sigma), 30 ng/ml progesterone (Sigma), 25 ng/ml triiodothyronine (Sigma) and 8 mg/ml putrescine (Sigma)]. Cells were either maintained in this medium where ‘non-myelinating’ conditions were required, or myelination was induced one week after Schwann cell addition by exposing the cells to myelination medium ((N2 medium, 1:300 phenol-free Matrigel (Corning), 5% charcoal-stripped FBS (ThermoFisher), 25 ng/ml recombinant- human NGF (Peprotech), 50mg/ml ascorbic acid (Sigma)). Myelinating co-cultures were maintained for a further 5 weeks before fixation for ICC analysis.

McDermott L.A., Weir G.A., Themistocleous A.C., Segerdahl A.R., Blesneac I., Baskozos G., Clark A.J., Millar V., Peck L.J., Ebner D., Tracey I., Serra J, & Bennett D.L. (2019). Defining the Functional Role of NaV1.7 in Human Nociception. Neuron, 101(5), 905-919.e8.

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Cultivation of Mouse Hepatocyte AML12 Cells

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Mouse hepatocyte AML12 cells were purchased from the Duke University Cell Culture Facility. AML12 cells were maintained in complete growth medium (CGM) which consisted of DMEM/F12 (US Biological) supplemented with 10% non-heat activated fetal bovine serum (FBS, GE Life Sciences Hyclone), penicillin and streptomycin (100 U/ml and 100 μg/ml, respectively, Gibco), 10 μg/ml insulin, 5 μg/ml transferrin, 7 ng/ml selenium (1:100 dilution of ITS, Millipore), and 100 nM dexamethasone (Sigma). Cells were grown in a humidified tissue culture incubator (Symphony, VWR) at 37 °C with 5% CO₂. For experiments, cells were harvested using trypsin (0.25%) –ethylenediaminetetraacetic acid and seeded into 96-well plates (5×10⁴ cells/well) in 200 μl of CGM and allowed to settle and grow for approximately 18 h at 37 °C with 5% CO₂.

Mello D.F., Trevisan R., Rivera N., Geitner N.K., Di Giulio R.T., Wiesner M.R., Hsu-Kim H, & Meyer J.N. (2019). Caveats to the use of MTT, Neutral Red, Hoechst and Resazurin to measure silver nanoparticle cytotoxicity. Chemico-biological interactions, 315, 108868.

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Chemicals Sourcing for Cell Experiments

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The chemicals used in this study were obtained from the following sources: FM 2–10, FM1-43, B-27 supplement and MEM (Invitrogen, Carlsbad, CA), Matrigel and FBS (Thermofisher, Waltham, MA) and transferrin (Millipore, Billerica, MA). All other chemicals were purchased from Sigma Aldrich.

Orock A., Logan S, & Deak F. (2017). Munc18-1 haploinsufficiency impairs learning and memory by reduced synaptic vesicular release in a model of Ohtahara syndrome. Molecular and cellular neurosciences, 88, 33-42.

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Generating Myelinating Co-Cultures from iPSC-derived Sensory Neurons

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Myelinating co-cultures were prepared as previously described.²⁴ (link)^,²⁵ (link) In brief, 30,000 rat Schwann cells were added to iPSCdSNs in Schwann cell basal medium [DMEM/F12 (ThermoFisher), 5 mg/ml insulin (Sigma), 100 mg/ml transferrin (Millipore), 25 ng/ml recombinant-human NGF (Peprotech), 25 ng/ml Selenium (Sigma), 25 ng/ml thyroxine (Sigma), 30 ng/ml progesterone (Sigma), 25 ng/ml triiodothyronine (Sigma) and 8 mg/ml putrescine (Sigma)]. Cells were maintained in this medium for one week to allow Schwann cell proliferation and alignment. Myelination was subsequently induced by exposing the cells to myelination medium ((N2 medium, 1:300 phenol-free Matrigel (Corning), 5% charcoal-stripped FBS (ThermoFisher), 25 ng/ml recombinant- human NGF (Peprotech), 50mg/ml ascorbic acid (Sigma)). Myelinating co-cultures were maintained for 8 or 26 weeks with twice weekly medium changes before fixation and immunocytochemistry.

Clark A.J., Kugathasan U., Baskozos G., Priestman D.A., Fugger N., Lone M.A., Othman A., Chu K.H., Blesneac I., Wilson E.R., Laurà M., Kalmar B., Greensmith L., Hornemann T., Platt F.M., Reilly M.M, & Bennett D.L. (2021). An iPSC model of hereditary sensory neuropathy-1 reveals L-serine-responsive deficits in neuronal ganglioside composition and axoglial interactions. Cell Reports Medicine, 2(7), 100345.

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