Corneal epithelial cells were cultured in Defined Keratinocyte-Serum Free medium (SFM) (Thermofisher Cat.No-10744019).The tissues collected from control patients was cultivated on type 1 collagen coated 1:10 Polydimethoxy siloxane (PDMS) of desired elastic modulus and 50 mg/ml of Gelatin Hydrogel in Defined Keratinocyte-SFM medium (Thermofisher Cat.No-10744019).Adherence of the tissues to culture plate was assured by a gravitational force from viscoelastic solution added on top of the tissues (HEALONOVD, Abbott Medical Optics, USA) as reported earlier44 (link),45 (link). After 24 h of incubation, tissues were fixed and immunofluorescence imaging was performed.
Defined keratinocyte sfm medium
Manufactured by Thermo Fisher Scientific
Sourced in United States
Defined Keratinocyte-SFM medium is a serum-free, defined culture medium designed for the growth and maintenance of human epidermal keratinocytes in vitro. It contains essential nutrients, growth factors, and other components required for the optimal growth of keratinocytes.
Automatically generated - may contain errors
Lab products found in correlation
Rpmi 1640 medium, by Thermo Fisher Scientific (3 mentions)
Lncap, by American Type Culture Collection (2 mentions)
Rwpe 1, by American Type Culture Collection (2 mentions)
Basic fibroblast growth factor (bfgf), by Merck Group (2 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (2 mentions)
Dmem f12 medium, by Thermo Fisher Scientific (2 mentions)
Insulin, by Merck Group (2 mentions)
Bovine serum albumin (bsa), by Merck Group (2 mentions)
Epidermal growth factor (egf), by Merck Group (2 mentions)
N2 nutrition, by STEMCELL (2 mentions)
Healon ovd, by Abbott (1 mentions)
Ham s f 12k medium, by Procell (1 mentions)
Rwpe 1, by Thermo Fisher Scientific (1 mentions)
Du145, by American Type Culture Collection (1 mentions)
Nt2 d1, by American Type Culture Collection (1 mentions)
Rpmi 1640, by Corning (1 mentions)
Lipofectamine 2000, by Thermo Fisher Scientific (1 mentions)
Trypsin edta, by Thermo Fisher Scientific (1 mentions)
Fetal bovine serum (fbs), by ExCell Bio (1 mentions)
10 protocols using defined keratinocyte sfm medium
1
Corneal Epithelial Cell Culture on Biomimetic Substrates
2
Cultivation and Manipulation of NPC Cell Lines
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Human NPC cell line HK1 was kindly gifted by Dr. Tao of the Chinese University of Hong Kong, and human NPC cell lines 5–8F, 6–10B, CNE1 and CNE2 and NP69, an immortalized normal human nasopharynx epithelial cell line, have been described previously by us33 (link),49 (link),50 (link). NPC cells were cultured with RPMI-1640(BI, Israel) medium supplemented with 10% fetal bovine serum (BI) at 37 °C in 5% CO2. NP69 cells were cultured with Defined Keratinocyte-SFM medium (Thermo Fischer Scientific) at 37 °C in 5% CO2. The cell lines were authenticated by short tandem repeat profiling prior to use, and were routinely tested negative for mycoplasma contamination using 4,6-diamidino-2-phenylindole staining.
Lentiviral interfering plasmid GV248-shHDAC7 and scramble nontarget shRNA control plasmid GV248-shNC, and a dual-luciferase reporter plasmid expressing wild-type EphA2 3′-UTR or mutant EphA2 3′-UTR in the predicted miR-4465 binding site were established by Genechem Inc. (Shanghai, China), and confirmed by sequencing. The core target sequence of shHDAC7 was 5′-GGCUGGAAACAGAAACCCA-3′. pENTER-HDAC7 expression plasmid and control plasmid were purchased from Vigene bioscience Inc. EphA2 expression plasmid and control plasmid have been described previously by us33 (link).
Lentiviral interfering plasmid GV248-shHDAC7 and scramble nontarget shRNA control plasmid GV248-shNC, and a dual-luciferase reporter plasmid expressing wild-type EphA2 3′-UTR or mutant EphA2 3′-UTR in the predicted miR-4465 binding site were established by Genechem Inc. (Shanghai, China), and confirmed by sequencing. The core target sequence of shHDAC7 was 5′-GGCUGGAAACAGAAACCCA-3′. pENTER-HDAC7 expression plasmid and control plasmid were purchased from Vigene bioscience Inc. EphA2 expression plasmid and control plasmid have been described previously by us33 (link).
3
Culture of Prostate Cell Lines
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The immortalized prostate luminal epithelial cell line RWPE1, and PCa cell lines PC3 and DU145, were acquired from ATCC. PC3 and DU145 cells were grown in Ham’s F-12K medium (Procell) and RPMI1640 medium (Invitrogen), respectively. RWPE1 cells were cultured in Defined Keratinocyte‐SFM medium (ThermoFisher). All medium was supplemented by 10% FBS and 1% P/S (penicillin/streptomycin); all cells were grown in 37°C with 5% CO2.
4
Cell Line Maintenance for Prostate Cancer Research
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Five cell lines, including a non–tumorigenic prostate epithelial cell (RWPE1), three PCa cells (LNCaP, 22RV1, and DU145), and one teratoma cell (NT2/D1), were originally purchased from the American Type Culture Collection and maintained in our laboratory. VCaP cell line was kindly gifted by Professor Qiao Zhou, Department of Pathology, West China Hospital, Sichuan University. RWPE1 cells were grown in Defined Keratinocyte‐SFM medium (ThermoFisher). LNCaP and 22RV1 cells were grown in RPMI‐1640 supplemented with 10% FBS. DU145, NT2/D1, and VCaP cells were maintained in DMEM supplemented with 10% FBS. All cells were incubated at 37°C with 5% CO2.
5
Culturing Prostate Cancer Cell Lines
6
Profiling Circular RNAs in Oral Squamous Cell Carcinoma
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HOK and CAL-27 cells were purchased from Tongpai Technology. SSC-9, HSC-3, and HSC-6 cells were obtained from CinoAsia Co., Ltd. HOK cells were cultured in defined Keratinocyte SFM medium (10744019, Thermo Fisher Scientific). CAL-27, HSC-3, and HSC-6 cells were appropriately cultured in DMEM (SH30243.01, Hyclone) supplemented with 10% FBS and 0.5% penicillin-streptavidin solution according to the vendor's guidelines. SSC-9 cells were cultured in DME/F-12 medium (SH30023.01, Hyclone) supplemented with 10% FBS and 0.5% penicillin-streptavidin mixture. Cells were kept in a humidified incubator at 37 C with 5% CO 2 . A total of 169 pairs of tumor and adjacent nontumor specimens were collected from clinically diagnosed patients with OSCC (97 males and 72 females, ages from 26 to 79 years old) in the Stomatological Hospital of Guangdong Province and immediately snap frozen in liquid nitrogen before undergoing further processing. Four pairs were deployed for circRNA microarray, which was performed on CapitalBio Technology Human CircRNA Array v2 chips (CapitalBio, Corp.) containing four identical arrays per slide. Each array possesses probes targeting 18 Â 10 4 human circRNA sequences from CircBase, Deepbase, and Rybak-Wolf 2015. Ten pairs were used to solidify the microarray results. All 169 pairs served evaluating correlation between circIGHG and OSCC prognosis.
7
Immortalization of Cyst Epithelial Cells
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Several single cysts 1–3 cm in diameter were dissected, sectioned with scissors, and then incubated with trypsin/EDTA (Gibco) at 37°C for 1 hour. Digested cyst cells were cultured in defined keratinocyte-SFM medium (Gibco) to select epithelial cells and exclude contaminating fibroblast cells. After epithelial cell selection for several weeks, RC cells were cultured and adapted in DMEM with 10% FBS at 37°C in 5% CO2.
For immortalization, primary cultured cyst cells were transfected with hTERT by Lipofectamine 2000 (Invitrogen). After transfection, cells were selected with 1 mg/ml G418 for two passages and then maintained with 0.5 mg/ml G418.
For immortalization, primary cultured cyst cells were transfected with hTERT by Lipofectamine 2000 (Invitrogen). After transfection, cells were selected with 1 mg/ml G418 for two passages and then maintained with 0.5 mg/ml G418.
8
Establishment and Characterization of Prostate Cancer Cell Lines
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The informed consent was obtained from the PCa patients and used for research only. This study was approved by the Ethics Committee of Shanghai Tenth People’s Hospital (No: SHSY-IEC-4.1/20-22/01), and the detailed information of the PCa patients was shown in Table S1 .
PC3, DU145, Lncap, 22RV1, and RWPE-1 cell lines were obtained from Shanghai Chinese Academy of Sciences (Shanghai, China). RWPE-1 cells were cultured in Defined Keratinocyte SFM medium (Gibco, USA), while PCa cell lines were cultivated with RPMI-1640 medium (Gibco) supplemented with 10% fetal bovine serum (FBS) (Gibco) and 1% penicillin/streptomycin (HyClone, Logan, UT, USA). PCSCs were grown in serum-free Dulbecco's modified Eagle's medium (DMEM)/F12 medium (Gibco) supplemented with 20 ng/ml epidermal growth factor (Sigma, St. Louis, USA), 20 ng/ml basic fibroblast growth factor (Sigma), 0.4% bovine serum albumin (BSA, Sigma), 5 μg/ml insulin (Sigma), and N2 nutrition (STEMCELL Technologies Inc., Canada) as previously described [28 (link)]. The cells were incubated at 37 °C in a humidified atmosphere of 5% CO2.
PC3, DU145, Lncap, 22RV1, and RWPE-1 cell lines were obtained from Shanghai Chinese Academy of Sciences (Shanghai, China). RWPE-1 cells were cultured in Defined Keratinocyte SFM medium (Gibco, USA), while PCa cell lines were cultivated with RPMI-1640 medium (Gibco) supplemented with 10% fetal bovine serum (FBS) (Gibco) and 1% penicillin/streptomycin (HyClone, Logan, UT, USA). PCSCs were grown in serum-free Dulbecco's modified Eagle's medium (DMEM)/F12 medium (Gibco) supplemented with 20 ng/ml epidermal growth factor (Sigma, St. Louis, USA), 20 ng/ml basic fibroblast growth factor (Sigma), 0.4% bovine serum albumin (BSA, Sigma), 5 μg/ml insulin (Sigma), and N2 nutrition (STEMCELL Technologies Inc., Canada) as previously described [28 (link)]. The cells were incubated at 37 °C in a humidified atmosphere of 5% CO2.
9
Establishment and Characterization of Prostate Cancer Cell Lines
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The informed consent was obtained from the PCa patients and used for research only. This study was approved by the Ethics Committee of Shanghai Tenth People’s Hospital (No: SHSY-IEC-4.1/20-22/01), and the detailed information of the PCa patients was shown in Table S1 .
PC3, DU145, Lncap, 22RV1, and RWPE-1 cell lines were obtained from Shanghai Chinese Academy of Sciences (Shanghai, China). RWPE-1 cells were cultured in Defined Keratinocyte SFM medium (Gibco, USA), while PCa cell lines were cultivated with RPMI-1640 medium (Gibco) supplemented with 10% fetal bovine serum (FBS) (Gibco) and 1% penicillin/streptomycin (HyClone, Logan, UT, USA). PCSCs were grown in serum-free Dulbecco's modified Eagle's medium (DMEM)/F12 medium (Gibco) supplemented with 20 ng/ml epidermal growth factor (Sigma, St. Louis, USA), 20 ng/ml basic fibroblast growth factor (Sigma), 0.4% bovine serum albumin (BSA, Sigma), 5 μg/ml insulin (Sigma), and N2 nutrition (STEMCELL Technologies Inc., Canada) as previously described [28 (link)]. The cells were incubated at 37 °C in a humidified atmosphere of 5% CO2.
PC3, DU145, Lncap, 22RV1, and RWPE-1 cell lines were obtained from Shanghai Chinese Academy of Sciences (Shanghai, China). RWPE-1 cells were cultured in Defined Keratinocyte SFM medium (Gibco, USA), while PCa cell lines were cultivated with RPMI-1640 medium (Gibco) supplemented with 10% fetal bovine serum (FBS) (Gibco) and 1% penicillin/streptomycin (HyClone, Logan, UT, USA). PCSCs were grown in serum-free Dulbecco's modified Eagle's medium (DMEM)/F12 medium (Gibco) supplemented with 20 ng/ml epidermal growth factor (Sigma, St. Louis, USA), 20 ng/ml basic fibroblast growth factor (Sigma), 0.4% bovine serum albumin (BSA, Sigma), 5 μg/ml insulin (Sigma), and N2 nutrition (STEMCELL Technologies Inc., Canada) as previously described [28 (link)]. The cells were incubated at 37 °C in a humidified atmosphere of 5% CO2.
10
Isolation and Cultivation of Human and Mouse Skin Cells
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Human keratinocytes (hKC) and fibroblasts (hFB) were isolated from foreskin following circumcision (age between 20 and 30 years) with ethical approval (No. 20191104-13) according to standard procedures [23 , 24 (link)]. Mouse keratinocytes (mKC) and fibroblasts (mFB) were isolated from neonatal dorsal skin as previously described [25 (link), 26 (link)]. hKC were cultivated in EpLife (Gibco, NY, USA) supplemented with EDGS as provided by the manufacturer. mKC were cultivated in Defined Keratinocyte-SFM medium (Gibco). Fibroblasts were cultivated in DMEM (GENOM) supplemented with 10% fetal bovine serum (ExCell bio, Shanghai, China). All cells were cultivated in an incubator with 5% CO2 at 37 °C.
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