Collagen coated plate

Manufactured by Corning

Sourced in United States

Collagen-coated plates are laboratory equipment designed for cell culture applications. The plates are coated with collagen, a naturally occurring extracellular matrix protein, to provide a suitable substrate for the attachment and growth of cells. These plates are intended to support the in vitro cultivation of various cell types.

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Lab products found in correlation

Glutamax, by Thermo Fisher Scientific (4 mentions) Percoll, by GE Healthcare (3 mentions) Pronase, by Roche (3 mentions) 100 mm cell vacuum strainer, by Merck Group (2 mentions) Epicult plus, by STEMCELL (2 mentions) Trypsin inhibitor, by Thermo Fisher Scientific (2 mentions) Insulin transferrin selenium, by Merck Group (2 mentions) Mgso4, by Thermo Fisher Scientific (2 mentions) Collagenase type 1, by Thermo Fisher Scientific (2 mentions) Collagenase type 2, by Thermo Fisher Scientific (2 mentions) William s e medium, by Lonza (2 mentions) Hepes, by Thermo Fisher Scientific (2 mentions) Dexamethasone (dex), by Merck Group (2 mentions) Cacl2 h2o, by Mallinckrodt (2 mentions) Pen strep, by Thermo Fisher Scientific (2 mentions) Ficoll paque plus, by GE Healthcare (1 mentions) Dmem f12 medium, by Welgene (1 mentions) Penicillin streptomycin, by Welgene (1 mentions) 4 oht, by Merck Group (1 mentions) Adriamycin, by Merck Group (1 mentions)

19 protocols using collagen coated plate

Isolation of Liver Cell Subpopulations

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Non-recirculating 2-step perfusion method with calcium and magnesium-free Hanks’ salt solution followed by a medium containing collagenase^{40 (link)} was used for the isolation of primary cells from liver tissues of C57BL/6 mice. The cell suspension was filtered through a 70 µm cell strainer and hepatocytes were obtained from the single cell-suspensions after 50 × g centrifugation. Hepatocytes with viability above 90% were plated on a collagen-coated plate (Corning, New York, USA).
The supernatant was centrifuged at 600 × g to obtain hepatic nonparenchymal cells. Primary HSCs were isolated by density-gradient centrifugation using ficoll paque-plus (GE Healthcare, Madison, WI) and percoll (GE Healthcare, Madison, WI, USA)^{41 (link)}. Non-parenchymal cell fraction was resuspended in a solution with 9 ml ficoll and 1 ml percoll. One milliliter PBS was added carefully and spin at 4 °C for 15 min at 1400 × g in centrifuge. HSC cell fraction was obtained from the layer between PBS and the ficoll/percoll gradient solution. Kupffer cells were isolated using Optiprep^{38 (link)}. The nonparenchymal cells are resuspended in 20% Optiprep. HBSS and 11.5% Optiprep were layered on the cell suspension and centrifuged at 1811 × g for 17 m. Kupffer cell fraction was obtained from the layer between 20% Optiprep and 11.5% Optiprep.

Kim H.Y., Choi Y.J., Kim S.K., Kim H., Jun D.W., Yoon K., Kim N., Hwang J., Kim Y.M., Lim S.C, & Kang K.W. (2021). Auranofin prevents liver fibrosis by system Xc-mediated inhibition of NLRP3 inflammasome. Communications Biology, 4, 824.

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Isolation and Culture of Murine Hepatocytes

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Primary hepatocytes were isolated from the liver of 8-week-old male mice by standard two-step collagenase perfusion and centrifugation. Isolated hepatocytes were maintained in William’s E media (#LM017-02; Welgene, Korea) on a collagen-coated plate (#354400; Corning, New York). AML12 cells were cultured in DMEM/F12 medium (#LM002-04; Welgene) with 10% FBS (#F2442; Sigma, Missouri) and 1% penicillin/streptomycin (#LS202-02; Welgene), supplemented with insulin (10 μg/ml), transferrin (5.5 μg/ml), selenium (5 ng/ml), and dexamethasone (40 ng/ml). HEK293, MIHA, and Huh7 cells were cultured in DMEM (#LM001-05; Welgene) containing 10% FBS and 1% penicillin/streptomycin. All cells were cultured at 37°C within a humidified chamber with 5% CO2.
YAP inhibitory signals were introduced as following: Serum starvation (16 h); high density (100% confluence, 24 h); sorbitol (0.5 M, 1 h); cytochalasin D (1 μM, 1 h); latrunculin B (0.25 mg/ml, 1 h); and 2-DG (2-deoxyglucose; 25 mM, 1 h). LB-100 was pretreated in LATS1/2 KO cells at a concentration of 5 μM for 1 h and in wild-type cells at a concentration of 10 μM for 1 h.

Kwon A., Lee N.Y., Yu J.H., Choi M.G., Park J, & Koo J.H. (2024). Mitochondrial stress activates YAP/TAZ through RhoA oxidation to promote liver injury. Cell Death & Disease, 15(1), 51.

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Mammary Gland Organoid Self-Renewal Assay

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Mammary glands were collected, cells isolated, and mammosphere cultures established as described previously (Cicalese et al., 2009 (link)). For organoid self-renewal assays, isolated MaProgs were cultured for 7 days in adhesion on collagen-coated plates (Corning) and, then, seeded at a density of 10,000 cells/well in 6-well ultralow attachment plates (Corning) in 5% matrigel (Corning) mammary colony medium (Panciera et al., 2016 (link)). Colonies were counted at the end of each passage (14 days), recovered in ice-cold HBSS, incubated with 400 μL Accutase (Sigma) for 5 minutes at 37°C and mechanically dissociated to obtain single-cell suspensions for serial replating.
Primary ErbB2-tumor cells were occasionally kept in short-term cultures in adhesion using DMEM/F12 (1:1, Lonza/GIBCO) supplemented with 10% FBS (HyClone, GE Healthcare Life Science), 10 mM HEPES (Sigma), 5 μg/mL insulin (Roche), 0.5 μg/mL hydrocortisone (Sigma), 10 ng/mL epidermal growth factor (EGF, Tebu-Bio), and 10 ng/mL cholera toxin (Sigma). In vitro treatments with Nut3 (Sigma, N6287), Adriamycin (Sigma, D1515) and 4-OHT (Sigma, H7904) were performed as indicated in the corresponding Results sections and Figures.

Santoro A., Vlachou T., Luzi L., Melloni G., Mazzarella L., D’Elia E., Aobuli X., Pasi C.E., Reavie L., Bonetti P., Punzi S., Casoli L., Sabò A., Moroni M.C., Dellino G.I., Amati B., Nicassio F., Lanfrancone L, & Pelicci P.G. (2019). p53 Loss in Breast Cancer Leads to Myc Activation, Increased Cell Plasticity, and Expression of a Mitotic Signature with Prognostic Value. Cell Reports, 26(3), 624-638.e8.

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Isolation of Bovine, Porcine, and Chicken MSCs

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Bovine top-round muscles, porcine hindshank, and chicken whole leg muscles were collected from female calves (17 weeks old), male porcine (3 days after birth), and embryonic chickens (16 days after fertilization), minced, digested with 1% pronase (Roche, Mannheim, Germany) for 1 h at 37 °C, and centrifuged at 1000×g for 3 min. Digested tissue was passed through a 100 mm cell vacuum strainer (Millipore, Darmstadt, Germany), and the filtrate was centrifuged at 1000×g for 5 min. Pellets were suspended in Ham's F-10 + 20% FBS +1% P&S + 5 ng/mL FGF2, seeded on collagen-coated plates (Corning, Brooklyn, NY, USA), and incubated in a 5% CO2 humidified incubator at 37 °C. The bovine, porcine, and chicken MSCs isolation method was adopted from our previous report (Lim et al., 2022 ).

Ahmad S.S., Lim J.H., Ahmad K., Chun H.J., Hur S.J., Lee E.J, & Choi I. (2024). Targeting myostatin using quercetin as a media supplement to improve myogenesis for cultured meat production: An in silico and in vitro study. Current Research in Food Science, 8, 100678.

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Sebocyte and ORS Cell Viability Assay

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Primary human sebocytes and ORS cells were plated in 96-well collagen-coated plates (CORNING, 5,000 cells per well) for 24 h. The day after seeding, sebocytes and ORS cells were incubated for 3 days without supplement medium in various concentrations of RG. MTT solution (3-[4,5] dimethylthiazol-2,5diphenyltetrazolium bromide) was then added at 70 μg per well for 3 hours. The formazan product was dissolved with DMSO, and optical density was measured at 570 nm.

Kwack M.H., & Lee W.J. (2021). Red Ginseng Water Extract Aggravates Inflammation in Sebocytes and Outer Root Sheath Cells After Treatment With Lipopolysaccharide and Mice With Cutibacterium Acnes-induced Inflammatory Nodules.

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Murine Mammary Organoid Culture

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The Trp53^tm1BrdBrca1^{tm1Aash/F22-24} Tg (BLG-cre) 74Acl/J mouse strain was purchased from The Jackson Laboratory (012620). All mice were maintained in a temperature-controlled room (21 °C) with a 12-h light-dark cycle with 40–60% humidity. Mammary glands from 6-week-old female mice were dissected out as organoids (around 1 cm³) and were cryopreserved in 10% DMSO/FBS in liquid nitrogen for long-term storage. Upon thawing, cryopreserved mammary organoids isolated from 6-week old mice were cut into small pieces and dissociated into single-cell suspensions with the gentleMACS dissociator (Miltenyi Biotec). After single-cell suspension was obtained, cells were cultured with EpiCult Plus (STEMCELL Technologies) on collagen-coated plates (Corning) and the medium was refreshed every other day. An aliquot of 5500 mammary cells was mixed with 50 μl Matrigel (Invitrogen) and 40 μl of the mixture was dispensed in the center of a well of a warm 24-well plate. The cell and Matrigel mixture solidified for 15 min at 37 °C. One ml of mouse mammary organoid growth medium (STEMCELL Technologies) was added to the well. Medium was refreshed 3 times a week.

He Z., Ghorayeb R., Tan S., Chen K., Lorentzian A.C., Bottyan J., Aalam S.M., Pujana M.A., Lange P.F., Kannan N., Eaves C.J, & Maxwell C.A. (2022). Pathogenic BRCA1 variants disrupt PLK1-regulation of mitotic spindle orientation. Nature Communications, 13, 2200.

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Isolation and Culture of Primary Hepatocytes and HSCs

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Primary hepatocytes and HSCs were isolated and cultured as previously described.²¹ (link) Briefly, mice were anesthetized with sodium pentobarbital (30 mg/kg intraperitoneally) and the portal vein was cannulated. The livers were perfused with ethylene glycol-bis(β-aminoethyl ether)-N,N,N′,N′-tetraacetic acid solution (5.4 mmol/L KCl, 0.44 mmol/L KH₂PO₄, 140 mmol/L NaCl, 0.34 mmol/L Na₂HPO₄, 0.5 mmol/L ethylene glycol-bis(β-aminoethyl ether)-N,N,N′,N′-tetraacetic acid, and 25 mmol/L Tricine, pH 7.2) followed by 0.075% type I collagenase (Sigma) in Gey's Balanced Salt Solution (Sigma). After an additional digestion step with 0.009% collagenase at 37°C agitation for 30 minutes, hepatocytes were separated with 25g centrifugation for 5 minutes at room temperature. The precipitated hepatocytes were cultured in Dulbecco's modified Eagle medium with 10% fetal bovine serum (FBS) on collagen-coated plates from Corning (Corning, NY). The supernatant was centrifuged further at 400g for 10 minutes at 4°C. The cell pellet was suspended in 11.5% OptiPrep and loaded carefully with Gey's Balanced Salt Solution. After centrifugation at 1400g for 20 minutes at 4°C, the interface fraction was collected and further washed with Gey's Balanced Salt Solution twice. The finally collected HSCs were cultured further in RPMI-1640 medium containing 10% FBS and 10% horse serum.

Zhou Z., Xu M.J., Cai Y., Wang W., Jiang J.X., Varga Z.V., Feng D., Pacher P., Kunos G., Torok N.J, & Gao B. (2018). Neutrophil–Hepatic Stellate Cell Interactions Promote Fibrosis in Experimental Steatohepatitis. Cellular and Molecular Gastroenterology and Hepatology, 5(3), 399-413.

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Isolation and Differentiation of Murine Muscle Stem Cells

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Gastrocnemius and cranial thigh muscles were collected from C57BL male mice (six weeks) and minced, digested with 1% pronase (Roche, Mannheim, Germany) for 1 h at 37 °C, and then centrifuged at 1000× g for 3 min followed by passage of the digested tissue phase through a 100 mm syringe filter (Millipore, Darmstadt, Germany). After centrifugation of the filtrate at 1000× g for 5 min, the pellets were suspended in DMEM + 20% FBS + 1% P/S + 5 ng/mL FGF2 (fibroblast growth factor 2, Miltenyi Biotec GmbH, Auburn, CA, USA), seeded on collagen-coated plates (Corning, Brooklyn, NY, USA), and incubated in a humidified 5% CO₂ atmosphere at 37 °C. The medium was changed every day. For induction of MSC differentiation into muscle cells, media were switched to DMEM + 2% FBS + 1% P/S or DMEM + 1% P/S followed by incubation for two days. MSC purity was confirmed with Pax7 protein expression (Santa Cruz Biotechnology, Paso Robles, CA, USA) using immunocytochemistry.

Lee E.J., Shaikh S., Choi D., Ahmad K., Baig M.H., Lim J.H., Lee Y.H., Park S.J., Kim Y.W., Park S.Y, & Choi I. (2019). Transthyretin Maintains Muscle Homeostasis through the Novel Shuttle Pathway of Thyroid Hormones during Myoblast Differentiation. Cells, 8(12), 1565.

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Neuronal Differentiation of SH-SY5Y Cells

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SH-SY5Y cells (ECACC, Sigma Aldrich, St. Louis, MO, USA) were cultured in a 1 : 1 mixture of Ham's F12 and Dulbeco's modified Eagle's medium (DMEM) (Gibco Life Technologies, Grand Island, NY, USA) supplemented with 10% heat-inactivated fetal bovine serum (FBS) (HyClone Labs., Logan, UT) and 100 U/mL of penicillin/streptomycin (Gibco Life Technologies, Grand Island, NY, USA) in a humidified atmosphere of 5% CO₂ at 37°C. The medium was changed twice a week, and cells were split at about 80% confluence. For neuronal differentiation, 5 × 10⁴ cells/well was seeded in collagen-coated plates (100 μg/mL, Corning, USA). After 24 hours (day 1), the medium was replaced by medium in which FBS concentration was reduced to 2% (differentiation medium) and supplemented with 10 μM all-trans retinoic acid (RA, Sigma Aldrich, Saint Louis, MO). Cells were incubated for 5 days, with the medium replaced every day, except on the second day. At day 5 of differentiation, the medium was removed, and cultures were stimulated with serum-free medium supplemented with human brain derived neurotrophic factor (BDNF 50 ng/mL—R&D Systems, MN, USA). At day 7 of differentiation, neurons were used in the experiments.

Arruda G.L., Vigerelli H., Bufalo M.C., Longato G.B., Veloso R.V., Zambelli V.O., Picolo G., Cury Y., Morandini A.C., Marques A.C, & Sciani J.M. (2021). Box Jellyfish (Cnidaria, Cubozoa) Extract Increases Neuron's Connection: A Possible Neuroprotector Effect. BioMed Research International, 2021, 8855248.

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Cryopreserved Mammary Epithelial Cell Isolation

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Highly purified MECs were isolated from cryopreserved mammary glands from 6-week-old mice. Briefly, this process involved dissociating viably cryopreserved preparations into single-cell suspensions. Cryopreserved mammary glands (around 1 cm³) were cut into small pieces for better tissue dissociation, which was performed with gentleMACS (Miltenyi Biotec) following the manufacturer’s protocols. After obtaining a single-cell suspension, cells were cultured with EpiCult Plus (STEMCELL Technologies) on collagen-coated plates (Corning) and the medium was refreshed every other day. For CFC assays, MECs were seeded at single-cell density (100 cells per 24 wells) in collagen-coated plates for 5 days, and LPS at different doses was used to activate NF-κB in MEC cultures.

Mateo F., He Z., Mei L., de Garibay G.R., Herranz C., García N., Lorentzian A., Baiges A., Blommaert E., Gómez A., Mirallas O., Garrido-Utrilla A., Palomero L., Espín R., Extremera A.I., Soler-Monsó M.T., Petit A., Li R., Brunet J., Chen K., Tan S., Eaves C.J., McCloskey C., Hakem R., Khokha R., Lange P.F., Lázaro C., Maxwell C.A, & Pujana M.A. (2022). Modification of BRCA1-associated breast cancer risk by HMMR overexpression. Nature Communications, 13, 1895.

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