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Pregm

Manufactured by Lonza
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PrEGM is a cell culture media designed for the growth and maintenance of primary human prostate epithelial cells. It provides the necessary nutrients and growth factors to support the proliferation and differentiation of these cells in vitro.

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Lab products found in correlation

Matrigel, by BD (7 mentions) Prebm, by Lonza (5 mentions) Matrigel, by Corning (4 mentions) Insulin transferrin selenium, by Thermo Fisher Scientific (4 mentions) Dispase, by Thermo Fisher Scientific (4 mentions) Dispase solution, by Thermo Fisher Scientific (2 mentions) Matrigel prostate epithelial growth medium, by Corning (2 mentions) Neutralized collagen, by BD (2 mentions) P160 rock inhibitor y 27632 dihydrochloride, by Selleck Chemicals (2 mentions) Purecol, by Advanced BioMatrix (2 mentions) Antibiotic antimycotic solution, by Thermo Fisher Scientific (2 mentions) Dulbecco s modified eagle s medium, by Thermo Fisher Scientific (2 mentions) Accutase, by STEMCELL (2 mentions) Mts assay, by Promega (2 mentions) Dispase, by STEMCELL (2 mentions) Trizol reagent, by Thermo Fisher Scientific (2 mentions) Y 27632, by Merck Group (2 mentions) T medium, by Thermo Fisher Scientific (1 mentions) Singlequots growth supplements, by Lonza (1 mentions) Foetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)

Close spelling variants of this product

PrEGM media (20 citations) PrEGM medium (8 citations) PrEGM Bullet kit (5 citations) PrEGM™ Prostate Epithelial Cell Growth Medium (2 citations)

40 protocols using pregm

1

Culturing Adult Prostate Cells

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Adult prostates from WT were harvested, dissociated and plated out in 96 well plates (104 cells per well), and 500ul of PrEGM (Lonza) was added. After 3 days, PrEGM was replaced with PrEBM (Lonza) containing hydrocortisone (5 μg/ml) and insulin/transferrin/selenium (Gibco) with or without Hgf (50 ng/ml). Cells were imaged after 3 days. To quantify the number of cells in each well, cells were trypsinized and counted using a haemocytometer.
Lim A., Shin K., Zhao C., Kawano S, & Beachy P.A. (2014). Spatially restricted Hedgehog signaling regulates HGF-induced branching of the adult prostate. Nature cell biology, 16(12), 1135-1145.
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2

Prostate Organoid Culture Protocol

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Adult prostates from WT or R26mTmG mice were harvested, dissociated, mixed with neutralized collagen (BD Biosciences), plated out in 48 well plates (5×104 cells per well), and 500 μl of PrEGM (Lonza) was added. Media was changed after 3 days. 5 days from the start of culture, PrEGM was replaced with PrEBM (Lonza) containing hydrocortisone (5 μg/ml) and insulin/transferrin/selenium (Gibco) with or without Hgf (50 ng/ml). Media was changed after 3 days and cells were imaged the following day. One representative experiment of three is presented.
Lim A., Shin K., Zhao C., Kawano S, & Beachy P.A. (2014). Spatially restricted Hedgehog signaling regulates HGF-induced branching of the adult prostate. Nature cell biology, 16(12), 1135-1145.
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3

Prostate Epithelial Cell Culture Optimization

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Either the bulk dissociated prostate epithelial cells or the FACS-purified basal and luminal cell populations were plated in T25 flasks precoated with PureCol (Advanced BioMatrix, San Diego, CA). We mainly used WIT medium (Stemgent, Cambridge, MA, cat no# 00-0045-500) supplemented with 10 μM of p160 ROCK inhibitor Y-27632 dihydrochloride (Selleckchem, Houston, TX) in this study. WIT Medium is a serum-free defined medium originally optimized for the robust culture of human primary mammary epithelial cells without the need of feeder cells57 (link). PrEGM (Prostate Epithelial Cell Growth Medium; Lonza, Walkersville, MD) has been widely used in culturing prostate cells in the field58 (link); however, we chose the WIT medium for most of our studies because we have observed that the WIT medium supports human primary prostate cells better than PrEGM. For cell passaging, the Trypsin-EDTA for Primary Cells (ATCC PCS-999-003) and Trypsin Neutralizing Solution (ATCC PCS-999-004) were utilized. In this study, freshly purified primary basal cells and short-term expanded cultures (in vitro and in vivo assays to characterize the epithelial biology.
Zhang D., Park D., Zhong Y., Lu Y., Rycaj K., Gong S., Chen X., Liu X., Chao H.P., Whitney P., Calhoun-Davis T., Takata Y., Shen J., Iyer V.R, & Tang D.G. (2016). Stem cell and neurogenic gene-expression profiles link prostate basal cells to aggressive prostate cancer. Nature Communications, 7, 10798.
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4

Prostate Sphere Assay Methodology

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The prostate sphere assay was performed as described previously (Xin et al., 2007 (link)). Briefly, 1×104 dissociated prostate cells were mixed in 1:1 Matrigel/PrEGM (Matrigel (BD Biosciences, San Jose, CA)/PrEGM (Lonza, Walkersville, MD)), plated in 12-well plates, and cultured in PrEGM medium. Prostate spheres were defined as spheroids with a diameter > 30 μm after a 6-day culture. When prostate basal cells were cocultured with stromal cells or urogenital sinus mesenchymal cells, the ratio of epithelial versus stromal cells is 1:2.
Wei X., Zhang L., Zhou Z., Kwon O.J., Zhang Y., Nguyen H., Dumpit R., True L., Nelson P., Dong B., Xue W., Birchmeier W., Taketo M., Xu F., Creighton C.J., Ittmann M.M, & Xin L. (2019). SPATIALLY-RESTRICTED STROMAL WNT SIGNALING RESTRAINS PROSTATE EPITHELIAL PROGENITOR PROLIFERATION THROUGH DIRECT AND INDIRECT MECHANISMS. Cell stem cell, 24(5), 753-768.e6.
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5

Prostate Organoid Culture Protocol

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Adult prostates from WT or R26mTmG mice were harvested, dissociated, mixed with neutralized collagen (BD Biosciences), plated out in 48 well plates (5×104 cells per well), and 500 μl of PrEGM (Lonza) was added. Media was changed after 3 days. 5 days from the start of culture, PrEGM was replaced with PrEBM (Lonza) containing hydrocortisone (5 μg/ml) and insulin/transferrin/selenium (Gibco) with or without Hgf (50 ng/ml). Media was changed after 3 days and cells were imaged the following day. One representative experiment of three is presented.
Lim A., Shin K., Zhao C., Kawano S, & Beachy P.A. (2014). Spatially restricted Hedgehog signaling regulates HGF-induced branching of the adult prostate. Nature cell biology, 16(12), 1135-1145.
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6

Culturing Adult Prostate Cells

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Adult prostates from WT were harvested, dissociated and plated out in 96 well plates (104 cells per well), and 500ul of PrEGM (Lonza) was added. After 3 days, PrEGM was replaced with PrEBM (Lonza) containing hydrocortisone (5 μg/ml) and insulin/transferrin/selenium (Gibco) with or without Hgf (50 ng/ml). Cells were imaged after 3 days. To quantify the number of cells in each well, cells were trypsinized and counted using a haemocytometer.
Lim A., Shin K., Zhao C., Kawano S, & Beachy P.A. (2014). Spatially restricted Hedgehog signaling regulates HGF-induced branching of the adult prostate. Nature cell biology, 16(12), 1135-1145.
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7

Culturing and Maintaining Cancer Cell Lines

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All cancer cell lines were purchased from ATCC (USA) and cultured in our lab following standard protocols. LNCaP human prostate cancer cells (ATCC #CRL-1740) were maintained in T-medium (Gibco, Formula no. 02-0056DJ) supplemented with heat-inactivated foetal bovine serum (10%, Gibco, 16000-044), L-Glutamine (2 mM) (Gibco, #25030-081) and Penicillin/Streptomycin (50 units/50 μg) (Gibco, #15070-063). MCF7 human breast cancer cells (ATCC #HTB-22) were maintained in RPMI-1640-based medium containing 5% (v/v) foetal bovine serum. PrEC normal human prostate epithelial cells (Cambrex Bio Science, #CC-2555, tissue acquisition number 13683) were maintained in prostate epithelial cell growth medium (PrEGM, Lonza, #CC-3165) supplemented with SingleQuots growth supplements (Lonza, #CC-4177).
Du Q., Bert S.A., Armstrong N.J., Caldon C.E., Song J.Z., Nair S.S., Gould C.M., Luu P.L., Peters T., Khoury A., Qu W., Zotenko E., Stirzaker C, & Clark S.J. (2019). Replication timing and epigenome remodelling are associated with the nature of chromosomal rearrangements in cancer. Nature Communications, 10, 416.
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8

Prostatosphere Assay for Prostate Stem Cells

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Prostatospheres were assayed as described (Lukacs et al., 2010a (link)). In brief,
3,000 Akt transformed prostate stem cells were resuspended in 100 μL of a 60:40 mixture of Matrigel (BD Bioscience,
NJ):PrEGM (Lonza, Basel, Switzerland) and plated in triplicate around the rim of a 12-well tissue culture plate. Matrigel was
allowed to solidify at 37°C, and 800 μL of PrEGM/well was added. Medium was changed twice weekly and spheres
were counted after 7 to 10 days. Each experiment was repeated 3 times. To passage the spheres, wells were treated with 1ml of
1 mg/ml Dispase solution (GIBCO, NJ). Spheres were digested with 0.25% trypsin with 2.21mM EDTA to obtain single cells, and
equal numbers of cells (2,000) were seeded in triplicate in Matrigel. The same protocol was used for all 6 passaged
generations.
Xiong X., Schober M., Tassone E., Khodadadi-Jamayran A., Sastre-Perona A., Zhou H., Tsirigos A., Shen S., Chang M., Melamed J., Ossowski L, & Wilson E.L. (2018). KLF4, A Gene Regulating Prostate Stem Cell Homeostasis, Is a Barrier to Malignant Progression and Predictor of Good Prognosis in Prostate Cancer. Cell reports, 25(11), 3006-3020.e7.
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9

Prostate Cancer Cell Line Cultivation

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CaP8 (Hong Wu, UCLA), MYC-CaP (Charles Sawyers, MSKCC), C42 (Leland Chung, Cedars Sinai), PEB-1 and PEL-1 (Lynette Wilson, NYU), LNCaP (ATCC), PC3 (ATCC), 293T (ATCC). Prostate cancer cell lines were grown in the following media conditions 5–10% (vol./vol.) FBS, 1% (vol./vol.) penicillin/streptomycin: RPMI-1640 for LNCaP, PC3 and C42. DMEM for 293T and MYC-CaP cells. PEB-1 and PEL-1 cells were grown in PrEGM (Lonza) with 1% (vol./vol.) penicillin/streptomycin and the addition of 10% only to PEL-1 cells.
Hu X., Garcia C., Fazli L., Gleave M., Vitek M.P., Jansen M., Christensen D, & Mulholland D.J. (2015). Inhibition of Pten deficient Castration Resistant Prostate Cancer by Targeting of the SET - PP2A Signaling axis. Scientific Reports, 5, 15182.
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10

Prostate Stem Cell Sphere Culture

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Sorted LSC population was re-suspended in 2:1 Matrigel/Prostate Epithelial growth medium (Matrigel, Cat#.354234, Corning; PrEGM, Cat#.CC-3166, Lonza) in a total volume of 120μL. Cells were seeded at a density of 8,000 or 10,000 cells per well in 12-well plates, and incubated for 7 days. Half medium change was performed every 2–3 days. Numbers of spheres were counted manually at 40x magnification at 7th day under microscope. To examine self-renewal ability of PSCs, spheres from primary culture were released by 1mg/mL Dispase (Cat#.17105-041, Invitrogen) at 37 °C for an hour, and dissociated into single cells for secondary sphere culture. To dissociate intact spheres, collected spheres were incubated with trypsin at room temperature (R.T.) for 10 mins. Trypsinized cells from spheres were passed through 26G needles three times. Dissociated cells were individually plated at 8,000 cells per well density in 12-well plates for 7 days. The numbers of secondary spheres were obtained at 7th day.
Wang H., Wang L., Jerde T.J., Chan B., Savran C.A., Burcham G.N., Crist S, & Ratliff T.L. (2015). Characterization of autoimmune inflammation induced prostate stem cell expansion. The Prostate, 75(14), 1620-1631.
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