60 mm cell culture dish

Manufactured by Corning

Sourced in United States

The 60-mm cell culture dish is a laboratory equipment used for the in vitro cultivation and growth of cells. It provides a suitable environment for cell attachment, proliferation, and maintenance. The dish has a diameter of 60 millimeters and is made of a high-quality material that is conducive for cell culture applications.

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Lab products found in correlation

Hypoxia incubator, by Thermo Fisher Scientific (1 mentions) Protease and phosphatase inhibitor cocktail, by Merck Group (1 mentions) Amersham ecl plus kit, by GE Healthcare (1 mentions) Ripa lysis buffer, by Santa Cruz Biotechnology (1 mentions) Pvdf membrane, by Bio-Rad (1 mentions) Dmem f12, by Thermo Fisher Scientific (1 mentions) Collagenase 1, by Merck Group (1 mentions) Gswp01300, by Merck Group (1 mentions) A1r confocal unit, by Nikon (1 mentions) Fetal calf serum (fcs), by Thermo Fisher Scientific (1 mentions) Collagenase type 1, by Merck Group (1 mentions) Ccl 2, by American Type Culture Collection (1 mentions) Penicillin streptomycin, by Corning (1 mentions) Fetal bovine serum (fbs), by Corning (1 mentions) Dulbecco modified eagle medium (dmem), by Corning (1 mentions)

7 protocols using 60 mm cell culture dish

Hypoxia and Insulin Effects on Pulmonary Endothelial Cells

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Experimental design No. 1: healthy pulmonary arterial endothelial cells (PAECs) at passages 2–4 were plated in a 60-mm cell culture dish (Corning) in 10% FBS M200 medium. After the cells grew to 60% confluence, a fresh medium with 10⁻⁷ mmol/L insulin was replaced. The dishes were then divided into normoxia and hypoxia groups and placed in a normoxia incubator (37°C, 210 ml/L O₂) and a hypoxia incubator (37°C, 100 ml/L O₂), respectively. The cells and culture media were collected after 6, 24, 48, and 96 h of incubation.
Experimental design No. 2: healthy PAECs at passages 2–4 were plated in a 60-mm cell culture dish (Corning) in 10% FBS M200 medium. For exposure to hypoxia, PAECs were placed in a hypoxia incubator (Thermo Fisher, Boston, MA, United States) filled with a mixture of 10% O2 and 5% CO2 at 37°C. The normoxic control group was filled with a mixture of 21% O2 and 5% CO2 at 37°C. Cells were treated with 10⁻⁷ mmol/L insulin. The cells were harvested after 48 h of incubation. The proteins were extracted for the immunoblotting test.

Fan F., He J., Su H., Zhang H., Wang H., Dong Q., Zeng M., Xing W, & Sun X. (2020). Tribbles Homolog 3-Mediated Vascular Insulin Resistance Contributes to Hypoxic Pulmonary Hypertension in Intermittent Hypoxia Rat Model. Frontiers in Physiology, 11, 542146.

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Stromal Fibroblast Signaling Pathway Analysis

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Stromal fibroblasts were seeded into a 60-mm cell culture dish (Corning) and proliferated to confluency. The culture medium was then changed to serum-free α-MEM with 0.3% bovine serum albumin, and stromal fibroblasts were cultured for 16 h. Culture was subsequently performed in serum-free α-MEM with 0.3% BSA with or without a sample or reagent. Stromal fibroblasts were placed on ice for 20 min in RIPA lysis buffer (50 mM Tris-HCl, 150 mM NaCl, 1% Nonidet P-40, 0.5% sodium deoxycholate, 0.1% SDS, pH 7.4; Santa Cruz Biotechnology, Dallas, TX, USA) containing protease and phosphatase inhibitor cocktails (both from Sigma-Aldrich) and lysed. The cell lysate was then collected and centrifuged at 13,400 × g at 4°C for 20 min; the supernatant was boiled in 5X SDS sample buffer and used in the following experiments. After performing electrophoresis on 20 μg of total cellular protein in a 10% SDS polyacrylamide gel, the gel was transferred to a PVDF membrane (Bio-Rad, Hercules, CA, USA). As primary antibodies, rabbit anti-phospho-Smad3 (diluted 1:2,000) and goat anti-human COX-2 antibodies were diluted with T-PBS and reacted at 4°C for 16 h. A luminescence reaction was performed with an Amersham ECL Plus kit (GE Healthcare, Uppsala, Sweden). Luminescent signals were detected using a Kodak Gel Logic 2200 Imaging System (Carestream, Rochester, NY, USA).

Yamada C., Aikawa T., Okuno E., Miyagawa K., Amano K., Takahata S., Kimata M., Okura M., Iida S, & Kogo M. (2016). TGF-β in jaw tumor fluids induces RANKL expression in stromal fibroblasts. International Journal of Oncology, 49(2), 499-508.

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Isolation and Characterization of Human ADSCs

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Human subcutaneous adipose tissue was obtained through liposuction with the patients’ written informed consent in the first affiliated hospital of Xi’an Jiaotong University. The human ethics committee approved the use of human-derived cells. Fresh isolated adipose tissue was cut into pieces, washed with phosphate buffered saline (PBS) and centrifuged at 1000 rpm for 5 min at room temperature. The lower part was discarded, and then the residual adipose tissue was digested with collagenase I (#SCR103, Sigma–Aldrich, St Louis, MI, USA) for 1 h at 37 ℃. The equivalent volume of complete Dulbecco’s modified Eagle’s medium F-12 (DMEM/F-12) (Gibco, Amarillo, TX, USA) was added into the tube to suspend digestion. Cells were collected after washing with PBS and then seeded in a 60 mm cell culture dish (Corning, NY, USA). Once reached confluence, adherent cells (passage 0) were digested with 0.125% trypsin-ethylenediaminetetraacetic acid (EDTA) and passaged at a 1:3 split ratio in 60 mm cell culture dishes. Cells of passage 5–10 were used in this study. ADSCs were stained with fluorescence-labelled monoclonal antibodies against CD29, CD44, CD90, CD105, CD45 and CD34 (eBioscience, San Diego, CA, USA) for flow cytometry (FCM) to analyze their surface phenotypes.

Shen Z., Wang W., Chen J., Chen B., Tang Y., Hou J., Li J., Liu S., Mei Y., Zhang L, & Lu S. (2022). Small extracellular vesicles of hypoxic endothelial cells regulate the therapeutic potential of adipose-derived mesenchymal stem cells via miR-486-5p/PTEN in a limb ischemia model. Journal of Nanobiotechnology, 20, 422.

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Live Imaging of Enteric Neural Crest Cells

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Live imaging of ENCCs was performed using a previously described suspended culture technique (Nishiyama et al., 2012 (link)). Briefly, E12.5 and E13.5 whole intestines from control (G4-RFP) and mutant (Spot^Tg/Tg;G4-RFP) embryos were placed on a small nitrocellulose filter (Millipore GSWP01300), which was then flipped on top of paraffin lines streaked in parallel on a 60-mm cell culture dish (Corning). Samples were cultured in Dulbecco's modified Eagle's medium (DMEM)/F12 medium containing 10% FBS and penicillin-streptomycin for 6 h (at 37°C and 5% CO₂) in a microscope incubation chamber (Okolab), and 250-μm-thick stacks were acquired every 10 min using a Nikon A1R confocal unit and a 10× objective. For calculations of speed and directionality of cell migration, 6-7 chain tip cells were tracked from at least three intestines of each genotype as previously described (Bergeron et al., 2015 (link); Soret et al., 2015 (link)). The net cell trajectory was determined by drawing a straight line relative to the longitudinal axis of the intestinal tissue from the cell position at 0 h and the radial position (in degrees) of the same cell at 6 h.

Bergeron K.F., Nguyen C.M., Cardinal T., Charrier B., Silversides D.W, & Pilon N. (2016). Upregulation of the Nr2f1-A830082K12Rik gene pair in murine neural crest cells results in a complex phenotype reminiscent of Waardenburg syndrome type 4. Disease Models & Mechanisms, 9(11), 1283-1293.

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Dental Mesenchymal Cell Isolation Protocol

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DMCs were isolated from the dental germs of 3-day-old WT and KO mice. Before surgery, the mice were sacrificed by cervical dislocation after anesthesia and sterilized using 75% ethanol for 5 min. Dental mesenchymal tissue was separated from the mandibular molar tooth germ by mechanical isolation, followed by digestion with type I collagenase (2mg/ml) (Sigma-Aldrich, MO, USA) for approximately 45min as previously described [18 (link)]. The tissue pieces were seeded into a 60mm cell culture dish (Corning, Tewksbury MA, USA), and cultured in Dulbecco’s modified Eagle’s medium (DMEM, Thermo Scientific, Waltham, USA), supplemented with 15% fetal calf serum (FCS, Gibco, Life Technologies, Carlsbad, CA, USA) and antibiotics (100 U/mL penicillin and 100 mg/mL streptomycin), grown in a humidified atmosphere at 37°C with 5% CO₂. Experiments were carried out using DMCs between the third and fifth passages.

Liao X., Feng B., Zhang D., Liu P., Zhou X., Li R, & Ye L. (2017). The Sirt6 gene: Does it play a role in tooth development?. PLoS ONE, 12(3), e0174255.

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Cell Cycle Synchronization and 9,10-PQ Treatment

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HeLa cells were purchased from the American Type Culture Collection (CCL-2; ATCC; Manassas, VA, USA). The cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM; Corning, NY, USA) and were supplemented with 10% fetal bovine serum (FBS; Corning) and 1% penicillin/streptomycin (GeneDepot; Barker, TX, USA). Cells were incubated in a humidified atmosphere containing 5% CO₂ at 37 °C. For cell cycle synchronization, 4 × 10⁵ cells were seeded in a 60 mm cell culture dish (Corning), and a double-thymidine (Thy-Thy) block was performed as previously described [38 (link)]. For chemical treatment, 2 × 10⁵ cells were seeded in each well of a 6-well plate or 35 mm confocal dish (SPL; Naechon, Pocheon,, Korea), to which was added a solution of 9,10-PQ (275034-25G, Sigma, Burlington, MA, USA) in dimethyl sulfoxide (DMSO) diluted in the medium to a concentration of 2.5 µM during incubation. Control cells were treated with an equivalent volume of DMSO. To eliminate ROS, 1 mM N-acetyl-L-cysteine (NAC; A8199) was added to the medium containing 2.5 µM 9,10-PQ, as previously described [39 (link)].

Kim S., Leem J., Oh J.S, & Kim J.S. (2022). Cytotoxicity of 9,10-Phenanthrenequinone Impairs Mitotic Progression and Spindle Assembly Independent of ROS Production in HeLa Cells. Toxics, 10(6), 327.

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Harvesting Multilayer Cell Sheets

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To harvest the multilayer cell sheets, we used a polyvinylidene difluoride (PVDF) membrane and forceps. The PVDF membranes were cut in a donut shape (PVDF, hydrophilic, 5.0 μm, 47 mm, white, plain, 100; Burlington, MA, USA) with the following size: 16 mm outer diameter and 9 mm inner diameter. The culture media was removed, and the cell sheets were washed with phosphate-buffered saline (PBS). The PBS was removed and replaced by fresh PBS. Forceps were used to detach the edges of the cells sheets and wrapped on the PVDF membrane placed in the center of the cell sheet. Once the edges were wrapped over the PVDF membrane, the whole PVDF/cell sheet was lifted with the forceps and placed in another 60 mm cell culture dish (Corning, Inc.), containing PBS. The cell sheets were unwrapped, and they were fixed in 10% Neutral buffered formalin for immunohistochemistry staining.

Oliva J., Florentino A., Bardag-Gorce F, & Niihara Y. (2019). Engineering, differentiation and harvesting of human adipose-derived stem cell multilayer cell sheets. Regenerative medicine, 14(3).

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