Murine epidermal growth factor

Manufactured by Merck Group

Murine epidermal growth factor is a laboratory product that functions as a cell growth factor derived from mice. It promotes the proliferation and differentiation of various cell types, including epidermal and epithelial cells.

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5 protocols using murine epidermal growth factor

Tumor Organoid Establishment from Mouse Models

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Firrm^−/−;Trp53^−/− and Trp53^−/− organoids were derived from cryopreserved mammary tumor pieces and processed as previously described by Duarte et al. (47 (link)). Briefly, tumor material was dissected and digested using collagenase A (2 mg/ml; Gibco) in advanced DMEM/F12, washed in growth medium (advanced DMEM/F12, 10 mM Hepes, GlutaMAX, and penicillin-streptomycin) and filtered through a cell strainer. Organoids were then cultured in growth medium further supplemented with B27 (Gibco), 125 μM N-acetyl-l-cysteine (Sigma-Aldrich), and murine epidermal growth factor (50 ng/ml; Sigma-Aldrich). Cells were embedded in 1:1 Culturex Reduced Growth Factor Basement Membrane Extract (BME) Type 2 (Trevigen) mixed with medium and maintained at 37°C and 5% CO₂. To establish organoid cultures, cells were supplemented with 5 μM nutlin (Sigma-Aldrich) for the initial 3 weeks of culture.

Mazouzi A., Moser S.C., Abascal F., van den Broek B., Del Castillo Velasco-Herrera M., van der Heijden I., Hekkelman M., Drenth A.P., van der Burg E., Kroese L.J., Jalink K., Adams D.J., Jonkers J, & Brummelkamp T.R. (2023). FIRRM/C1orf112 mediates resolution of homologous recombination intermediates in response to DNA interstrand crosslinks. Science Advances, 9(22), eadf4409.

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Establishment of Tumor-Derived Organoids

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ORG-KB1P4N.1 tumor-derived organoids were previously established.⁶¹ (link) Cultures were embedded in Culturex Reduced Growth Factor Basement Membrane Extract Type 2 (BME, Trevigen; 40 mL BME:growth media 1:1 drop in a single well of 24-well suspension plate) and grown in Advanced DMEM/F12 (GIBCO) supplemented with 1M HEPES (GIBCO), GlutaMAX (GIBCO), 50 units/ml penicillin-streptomycin (GIBCO), B27 (GIBCO), 125 mM N-acetyl-L-cysteine (Sigma) and 50 ng/mL murine epidermal growth factor (Sigma). Organoids were cultured under standard conditions (37°C, 5% CO2).

Bhin J., Paes Dias M., Gogola E., Rolfs F., Piersma S.R., de Bruijn R., de Ruiter J.R., van den Broek B., Duarte A.A., Sol W., van der Heijden I., Andronikou C., Kaiponen T.S., Bakker L., Lieftink C., Morris B., Beijersbergen R.L., van de Ven M., Jimenez C.R., Wessels L.F., Rottenberg S, & Jonkers J. (2023). Multi-omics analysis reveals distinct non-reversion mechanisms of PARPi resistance in BRCA1- versus BRCA2-deficient mammary tumors. Cell Reports, 42(5), 112538.

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Murine and Human Cell Line Cultures

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KB1P (KB1P-G3)³⁸ (link) and KB2P (KB2P-3.4)⁸⁹ (link) mouse tumor-derived cell lines were previously established and were grown in DMEM/F12 (Gibco) supplemented with 10% FBS and 50 units/ml penicillin-streptomycin (Gibco), containing 5 μg/mL Insulin (Sigma), 5 ng/mL cholera toxin (Sigma) and 5 ng/mL murine epidermal growth factor (Sigma), under low oxygen conditions (3% O2, 5% CO₂ at 37°C). RPE1-hTERT BRCA1^−/−;TRP53^−/− human cell line has been described before⁵⁷ (link) and was grown in DMEM+GlutaMAX (Gibco) supplemented with 10% FBS and 50 units/ml penicillin-streptomycin (Gibco), under low oxygen conditions (3% O2, 5% CO₂ at 37°C). SUM149PT (RRID:CVCL_3422) human cell line was grown in RPMI1640 (Gibco) medium supplied with 10% FBS and 50 units/ml penicillin-streptomycin (Gibco), under normal oxygen conditions (21% O2, 5% CO₂, 37°C).

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Establishment of Murine Mammary Tumor Organoids

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All lines have been described before (Duarte et al., 2018 (link)). ORG-KB1P.S and ORG-KB1P.R tumor organoids were derived from a PARPi-naive and PARPi-resistant K14cre;Brca1^F/F/;Trp53^F/F (KB1P) mouse mammary tumor, respectively. The ORG-KP tumor organoid line was derived from a K14cre;Trp53^F/F;Abcb1a^−/−;Abcb1b^−/− (KPM) mouse mammary tumor. Cultures were embedded in Culturex Reduced Growth Factor Basement Membrane Extract Type 2 (BME, Trevigen; 40 mL BME:growth media 1:1 drop in a single well of 24-well plate) and grown in Advanced DMEM/F12 (GIBCO) supplemented with 1M HEPES (GIBCO), GlutaMAX (GIBCO), 50 units/ml penicillin-streptomycin (GIBCO), B27 (GIBCO), 125 mM N-acetyl-L-cysteine (Sigma) and 50 ng/ml murine epidermal growth factor (Sigma). Organoids were cultured under standard conditions (37°C, 5% CO₂) and regularly tested for mycoplasma contamination.

Dias M.P., Tripathi V., van der Heijden I., Cong K., Manolika E.M., Bhin J., Gogola E., Galanos P., Annunziato S., Lieftink C., Andújar-Sánchez M., Chakrabarty S., Smith G.C., van de Ven M., Beijersbergen R.L., Bartkova J., Rottenberg S., Cantor S., Bartek J., Chaudhuri A.R, & Jonkers J. (2021). Loss of nuclear DNA ligase III reverts PARP inhibitor resistance in BRCA1/53BP1 double-deficient cells by exposing ssDNA gaps. Molecular cell, 81(22), 4692-4708.e9.

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3D Mammary Tumor Organoid Culture

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The KB1P4 3D tumor organoid line was previously established from a Brca1 À/À ;p53 À/À mouse mammary tumor and cultured as described (Duarte et al., 2018) . Briefly, cultures were embedded in Culturex Reduced Growth Factor Basement Membrane Extract Type 2 (BME, Trevigen; 40 mL BME:growth media 1:1 drop in a single well of 24-well plate) and grown in Advanced DMEM/F12 (AdD-MEM/F12, GIBCO) supplemented with 1 M HEPES (Sigma), GlutaMAX (GIBCO) 50 U/ml penicillin-streptomycin (GIBCO), B27 (GIBCO), 125 mM N-acetyl-L-cysteine (Sigma) and 50 ng/ml murine epidermal growth factor (Sigma). Organoids were cultured under standard conditions (37 C, 5% CO 2 ) and regularly tested for mycoplasma contamination.
Further in vitro culture details and gene editing details are provided in the Method Details section.

Francica P., Mutlu M., Blomen V.A., Oliveira C., Nowicka Z., Trenner A., Gerhards N.M., Bouwman P., Stickel E., Hekkelman M.L., Lingg L., Klebic I., van de Ven M., de Korte-Grimmerink R., Howald D., Jonkers J., Sartori A.A., Fendler W., Chapman J.R., Brummelkamp T, & Rottenberg S. (2020). Functional Radiogenetic Profiling Implicates ERCC6L2 in Non-homologous End Joining. Cell reports, 32(8).

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