Basic fibroblast growth factor (bfgf)

Manufactured by Bio-Techne

Sourced in Portugal

BFGF is a recombinant human basic Fibroblast Growth Factor (bFGF) produced in E. coli. bFGF is a heparin-binding growth factor that stimulates the proliferation of a variety of cell types, including fibroblasts, myoblasts, osteoblasts, and vascular endothelial cells.

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Lab products found in correlation

Epidermal growth factor (egf), by Bio-Techne (2 mentions) Insulin, by Merck Group (2 mentions) Te2000 u microscope, by Nikon (2 mentions) Sp600125, by Bio-Techne (1 mentions) Pd0325901, by Cayman Chemical (1 mentions) High glucose dmem f12, by Thermo Fisher Scientific (1 mentions) Forskolin, by Merck Group (1 mentions) Albumax 1, by Thermo Fisher Scientific (1 mentions) Sb203580, by Bio-Techne (1 mentions) N2 supplement, by Thermo Fisher Scientific (1 mentions) Basic fibroblast growth factor (bfgf), by Merck Group (1 mentions) Tgf β, by Thermo Fisher Scientific (1 mentions) Ascorbic acid, by Merck Group (1 mentions) Chir99021, by Axon Medchem (1 mentions) Dorsomorphin, by Bio-Techne (1 mentions) Dmem f12, by Corning (1 mentions) Rock inhibitor, by Bio-Techne (1 mentions) Basic fibroblast growth factor (bfgf), by Thermo Fisher Scientific (1 mentions) Glutamax, by Thermo Fisher Scientific (1 mentions) N2 neuroplex, by Gemini Bio (1 mentions)

5 protocols using basic fibroblast growth factor (bfgf)

Naive Conversion of Primed hESCs

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Three primed hESC lines were subjected to naive conversion on MEFs in low oxygen conditions in either: (i) naive human stem cell medium (NHSM) composed of basal hESC media supplemented with AlbumaxI, N2 supplement (Invitrogen), PD0325901 (1 μM, Cayman) and CHIR99021 (3 μM, Axon Medchem; known together as 2i), leukaemia inhibitory factor (LIF, 1000U, Sigma), bFGF, (8 ng ml⁻¹), SP600125 (10 μM, Tocris), SB203580 (10 μM, Tocris) and TGFβ (1 ng ml⁻¹, Peprotech)10 (link); (ii) RT consisting of high glucose DMEM-F12 (Invitrogen) further supplemented with histone deacetylase inhibitors, sodium butyrate (0.1 mM) and SAHA (50 nM) for first three passages followed by 2i and bFGF (10 ng ml⁻¹)11 (link) or (iii) our novel medium naive conversion medium (NCM) containing hESC media with added 2i, LIF, forskolin (10 μM, Sigma), ascorbic acid (50 ng ml⁻¹, Sigma) and bFGF(12 ng ml⁻¹)8 (link). Upon the appearance of domed-shaped colonies, the culture was passaged using 0.05% trypsin and once the culture was well established, cells were split at a ratio of 1:4 to 1:8 every 3 days. Single-cell clonogenicity was determined from the number of single-cells plated (upon dissociating primed and naive hESCs using trypsin without ROCKi) and the subsequent number of colonies formed. Doubling time was calculated using the formula:

Warrier S., Van der Jeught M., Duggal G., Tilleman L., Sutherland E., Taelman J., Popovic M., Lierman S., Chuva De Sousa Lopes S., Van Soom A., Peelman L., Van Nieuwerburgh F., De Coninck D.I., Menten B., Mestdagh P., Van de Sompele J., Deforce D., De Sutter P, & Heindryckx B. (2017). Direct comparison of distinct naive pluripotent states in human embryonic stem cells. Nature Communications, 8, 15055.

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Neuronal Differentiation from iPSCs

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Neural differentiation was performed as described elsewhere (Thomas et al, 2017). Briefly, iPSC media was replaced with DMEM/F12 (Corning Cellgro) with 1× HEPES, 1× penicillin‐streptomycin, 1× Glutamax (Life Technologies) and 1× N2 NeuroPlex (Gemini Bio‐products), supplemented with 1 mM dorsomorphin (Tocris) and 10 mM SB431542 (StemGent). Next, colonies were kept in suspension for 7 days to form embryoid bodies (EBs), which were plated onto matrigel‐coated plates and cultured using DMEM/F12 with 1× HEPES, 1× penicillin‐streptomycin, Glutamax, 0.5× N2 NeuroPlex and 1× Gem21 NeuroPlex (Gemini Bio‐products), supplemented with 20 ng/ml bFGF (Life Technologies). Neural rosettes were manually collected, gently dissociated with accutase (Stem Cell Technologies), and the neural progenitor cells (NPCs) were re‐plated on poly‐L‐ornithine/laminin‐coated plates. Next, the NPCs were differentiated into neurons by adding of 5 μM ROCK inhibitor (Tocris) for 48 h and bFGF withdraw for 6 weeks.

Baltussen L.L., Negraes P.D., Silvestre M., Claxton S., Moeskops M., Christodoulou E., Flynn H.R., Snijders A.P., Muotri A.R, & Ultanir S.K. (2018). Chemical genetic identification of CDKL5 substrates reveals its role in neuronal microtubule dynamics. The EMBO Journal, 37(24), e99763.

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Hematopoietic Differentiation of hiPSCs

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We performed hematopoietic differentiation of hiPSCs using a modified version of the EB-based hematopoietic differentiation protocol described previously (Lachmann et al., 2014; van Wilgenburg et al., 2013 ). In brief, PSC colonies were disrupted to fragments using collagenase-V, and EB formation was induced by cultivation for 5 days in ESC medium supplemented with 10 ng/mL bFGF and 10 μM Rock inhibitor (Y-27632; Tocris) in six-well suspension plates on an orbital shaker (100 rpm). After manual transfer of EBs onto tissue culture six-well plates and cultivation in differentiation medium I (APEL medium; Stem Cell Technologies) supplemented with 1% penicillin-streptomycin (Life Technologies), 25 ng/ml human IL-3 (hIL-3) and 50 ng/ml human M-CSF (hM-CSF), human G-CSF (hG-CSF), or human GM-CSF (hGM-CSF; all from Peprotech), MCFCs were generated from the attached EBs within 7 days. From d10–d15 onward, monocytes/macrophages or granulocytes generated by the MCFCs were harvested twice a week from the supernatant and filtered through a 150-mM mesh. For further maturation, monocytes/macrophages or granulocytes were cultured in differentiation medium II (RPMI1640 medium supplemented with 10% fetal serum, 2 mM L-glutamine, 1% penicillin-streptomycin, and 100 ng/ml hM-CSF, hG-CSF, or hGM-CSF, respectively) for 7–10 days.

Lachmann N., Ackermann M., Frenzel E., Liebhaber S., Brennig S., Happle C., Hoffmann D., Klimenkova O., Lüttge D., Buchegger T., Kühnel M.P., Schambach A., Janciauskiene S., Figueiredo C., Hansen G., Skokowa J, & Moritz T. (2015). Large-Scale Hematopoietic Differentiation of Human Induced Pluripotent Stem Cells Provides Granulocytes or Macrophages for Cell Replacement Therapies. Stem Cell Reports, 4(2), 282-296.

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HCT116 Cell Spheroid Formation and Treatment

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HCT116 cells were resuspended in serum-free stem cell culture media consisting of Dulbecco’s modified Eagle medium supplemented with 10 ng/mL bFGF and 20 ng/mL EGF (Bio-techne, Citomed Lda, Lisboa, Portugal), 1× B27 (Life Technologies), and 5 μg/mL insulin (Sigma-Aldrich). Cells were plated in 24-well ultra-low attachment plates (Corning Inc.; one spheroid/well) at 1 × 10³ cells/well. Treatments with Roy-Bz or vehicle were performed 3 days after spheroid formation for up to 96 h, or at the seeding time for 48 h. Spheroid formation was monitored using an inverted Nikon TE 2000-U microscope at ×100 magnification with a DXM1200F digital camera and using Nikon ACT-1 software. Spheroid diameters were quantified using the ImageJ software.

Bessa C., Soares J., Raimundo L., Loureiro J.B., Gomes C., Reis F., Soares M.L., Santos D., Dureja C., Chaudhuri S.R., Lopez-Haber C., Kazanietz M.G., Gonçalves J., Simões M.F., Rijo P, & Saraiva L. (2018). Discovery of a small-molecule protein kinase Cδ-selective activator with promising application in colon cancer therapy. Cell Death & Disease, 9(2), 23.

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Mammosphere Assay with BBIT20, CDDP and Olaparib

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A total of 1.5 × 10³ HCC1937 cells/well was seeded in 24-well plates covered with 1% agarose in DMEM:F12 supplemented with 20 ng ml⁻¹ bFGF, 40 ng ml⁻¹ EGF (Bio-techne, Citomed Lda, Lisboa, Portugal), 1 × B27 (Life Technologies, Porto, Portugal), 10 μg ml⁻¹ insulin (Sigma-Aldrich, Sintra, Portugal) and 2 mM L-Glutamine (Sigma) and treated with BBIT20 at the seeding time (Bessa et al., 2018 (link)). Mammospheres also grown in compound-free medium for 3 followed by BBIT20 treatment (Bessa et al., 2018 (link)).
Using 3-day-old mammospheres, BBIT20 synergistic potential was evaluated through analysis of its effect, alone and in combination with cisplatin (CDDP) or olaparib, on spheroid growth for additional 14 days. New medium with the drugs (or vehicle only) was added to the wells at days 1, 3, 6, 9 and 12. Mammospheres were monitored using an inverted Nikon TE 2000-U microscope at 100X magnification, with DXM1200F digital camera and Nikon ACT-1 software (Raimundo et al., 2018 (link)). Spheroid diameters were quantified using Fiji Software (Schindelin et al., 2012 (link)).

Raimundo L., Paterna A., Calheiros J., Ribeiro J., Cardoso D.S., Piga I., Neto S.J., Hegan D., Glazer P.M., Indraccolo S., Mulhovo S., Costa J.L., Ferreira M.J, & Saraiva L. (2021). BBIT20 inhibits homologous DNA repair with disruption of the BRCA1–BARD1 interaction in breast and ovarian cancer. British journal of pharmacology, 178(18), 3627-3647.

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