Fish collagen type I (FCOL) was purchased from Creative Enzymes Inc. (Shirley, NY, USA), hyaluronic acid (HA) was purchased from Wisapple Biotech Co, Ltd. (Beijing, China), Pronatal® LF 10/60 SA (SA 10/60) with G/M % ratios of 70/30 was kindly gifted by IMCD UK Limited, (Surrey, UK), sodium alginate (SA) with G/M % ratio 39/61, Gelatin (GEL) and D-mannitol (D-mann) were purchased from Sigma-Aldrich (Gillingham, UK)., bovine serum albumin (BSA), and calcium chloride were obtained from Acros Organics (Branchburg, NJ, USA) while sodium chloride (NaCl) was purchased from Fisher Scientific, (Loughborough, UK). Adult human primary epidermal keratinocytes [PCS-200-011, ATCC], human dermal fibroblasts [PCS-200-011, ATCC], dermal cell basal medium [PCS-200-030, ATCC], keratinocytes growth kit [PCS-200-040, ATCC], ATCC and Dulbecco’s Modified Eagle’s Medium (DMEM) [PCS-200-030, ATCC] were purchased from LGC standards (Middlesex, UK). Methyl thiazolyldiphenyl-tetrazolium bromide (MTT) and trypan blue stain were obtained from Thermo Fisher Scientific (Paisley, UK), fetal bovine serum was purchased from Sigma Aldrich, (Dorset, UK).
Pcs 200 030
Manufactured by American Type Culture Collection
Sourced in United Kingdom
The PCS-200-030 is a laboratory equipment product offered by American Type Culture Collection. It serves as a core function of cell culture processing.
Automatically generated - may contain errors
Lab products found in correlation
Fetal bovine serum (fbs), by Thermo Fisher Scientific (4 mentions)
Pcs 200 040, by American Type Culture Collection (3 mentions)
Keratinocyte growth kit, by American Type Culture Collection (3 mentions)
Pcs 200 013, by American Type Culture Collection (3 mentions)
Pcs 200 011, by American Type Culture Collection (2 mentions)
Fetal bovine serum (fbs), by Merck Group (2 mentions)
Pcs 200 014, by American Type Culture Collection (2 mentions)
Pcs 200 042, by American Type Culture Collection (2 mentions)
Ps 200 010, by American Type Culture Collection (2 mentions)
Penicillin, by Thermo Fisher Scientific (2 mentions)
Sk mel 3, by American Type Culture Collection (2 mentions)
A2058, by American Type Culture Collection (2 mentions)
Trypan blue stain, by Thermo Fisher Scientific (1 mentions)
Calcium chloride, by Thermo Fisher Scientific (1 mentions)
Sodium alginate, by Merck Group (1 mentions)
Gelatin gel, by Merck Group (1 mentions)
Bovine serum albumin (bsa), by Thermo Fisher Scientific (1 mentions)
Sodium chloride (nacl), by Thermo Fisher Scientific (1 mentions)
Dulbecco s modified eagle s medium dmem, by American Type Culture Collection (1 mentions)
Methyl thiazolyldiphenyl tetrazolium bromide mtt, by Thermo Fisher Scientific (1 mentions)
12 protocols using pcs 200 030
1
Collagen-Hyaluronic Acid Biomaterial Formulation
2
Culturing HGECs and BMMCs for P. gingivalis stimulation
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HGECs were cultured according to the manufacturer’s protocol (PCS-200-014, American Type Culture Collection (ATCC), Manassas, VA, USA). The cells were sub-cultured in Dermal Cell Basal Medium (PCS-200-030, ATCC, Manassas, VA, USA) supplemented with Keratinocyte Growth Kit (PCS-200-040, ATCC, Manassas, VA, USA) and 100 U penicillin/streptomycin (P4333-100ML, Sigma-Aldrich, St. Louis, MO, USA) at 37 °C in humidified air with 5% CO2. A confluent culture of HGECs was stimulated with P. gingivalis (10% formalin-fixed, strain 3327, ATCC, Manassas, VA, USA) (107 colony-forming units/mL) for 12 h to examine mRNA expression. As a negative control, HGECs were cultured without bacterial stimulation. P. gingivalis treatment was performed as previously described [20 (link)].
Bone marrow mononuclear cells (BMMCs) were collected from the femurs and tibias of C57BL/6J Jcl mice by density gradient centrifugation with Histopaque-1083 (Sigma-Aldrich, St. Louis, MO, USA) in complete alpha-modified Eagle’s minimum essential medium (Sigma-Aldrich), containing 10% fetal bovine serum (Invitrogen, Carlsbad, CA, USA), L-glutamine (Sigma-Aldrich, St. Louis, MO, USA), and antibiotics (penicillin, streptomycin, and gentamicin; Invitrogen, Carlsbad, CA, USA).
Bone marrow mononuclear cells (BMMCs) were collected from the femurs and tibias of C57BL/6J Jcl mice by density gradient centrifugation with Histopaque-1083 (Sigma-Aldrich, St. Louis, MO, USA) in complete alpha-modified Eagle’s minimum essential medium (Sigma-Aldrich), containing 10% fetal bovine serum (Invitrogen, Carlsbad, CA, USA), L-glutamine (Sigma-Aldrich, St. Louis, MO, USA), and antibiotics (penicillin, streptomycin, and gentamicin; Invitrogen, Carlsbad, CA, USA).
3
Establishment of Oral Cancer Cell Lines
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Human OSCC cell lines derived from oral cancer (SAS, HSC-2, HSC-3, Ca9-22, OSC-19, OSC-20, SAT, and KON) were purchased from the National Institute of Biomedical Innovation (Osaka, Japan). The HOC-31354 (link) and TSU55 (link) cell lines were kindly provided by Professor Kawashiri (Kanazawa University). HNOK cells were purchased from the American Type Culture Collection (ATCC; Manassas, VA, USA; PCS-200-014). SAS-R and HSC-2-R, which were established from SAS and HSC-2 cells, were used as the CRR cell lines. The CRR cell lines were produced by exposing cells to gradually increasing X-ray doses.5 (link) The OSCC cell lines were cultured in Dulbecco’s modified Eagle medium (DMEM; D6429; Sigma-Aldrich, Saint Louis, MO, USA) supplemented with 10% fetal bovine serum (Sigma-Aldrich) at 37°C and 5% CO2. HNOK cells were cultured in dermal cell basal medium (PCS-200-030; ATCC) supplemented with the Keratinocyte Growth Kit (PCS-200-040; ATCC). CRR cells continued to proliferate under a daily IR dose of 2 Gy for >30 days in vitro.
4
Diverse Mammalian Cell Lines for Research
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Following mammalian cell lines were used as the test models: epithelial cells of malignant melanoma A375 (ATCC® CRL1619™), epithelial cells of colorectal adenocarcinoma Caco-2 (ATCC® HTB37™), primary colon fibroblasts CCD112CoN (ATCC® CRL1541™) and primary epidermal melanocytes (PEM) (ATCC® PCS200013™). The cell lines used were purchased from ATCC® (Manassas, VA, US). All the cells, except primary melanocytes, were grown in Dulbecco’s modified Eagle’s Medium (DMEM) (Sigma, St. Louis, MO, US), supplemented with 10% fetal bovine serum (FBS) and 2 mM glutamine at 37 °C and 5% CO2. Primary melanocytes were cultured in Dermal Cell Basal Medium (ATCC® PCS200030) supplemented with Adult Melanocyte Growth Kit (ATCC® PCS200042).
5
Culturing Melanoma Cell Lines and Melanocytes
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Human melanoma cell line A375 (malignant, CRL-1619), G361 (malignant, CRL-1424), A2058 (metastatic, CRL-11147), SK-MEL-3 (metastatic, HTB-69), normal human primary epidermal melanocytes (PCS-200-013) were all purchased from American Type Culture Collection (ATCC). A375, A2058 cell lines were cultured in DMEM (ATCC) supplemented with 10% fetal bovine serum (FBS, Gibco). G361, SK-MEL-3 cell lines were cultured in McCoy's 5A (ATCC) supplemented with 10% and 15% fetal bovine serum (FBS, Gibco), respectively. Normal human primary epidermal melanocytes were grown in dermal cell basal media (ATCC, PCS-200-030) supplemented with adult melanocyte growth kit (ATCC, PCS-200-042) components. All cells were grown in a humidified incubator at 37°C, 5% CO2 atmosphere. After 1-2 days, cells were chosen to perform calcium imaging or other functional experiments.
6
Keratinocyte TWEAK and Cytokine Stimulation
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The mouse keratinocyte cell line PAM212 (generous gift from Wendy Havran, Scripps Institute, La Jolla, CA) and normal human primary epidermal keratinocytes from neonates (NHEK, PS-200-010, ATCC, Manassas, VA) were grown in 10% fetal calf serum-supplemented DMEM medium and Dermal Cell Basal Medium (PCS-200-030, ATCC), respectively. Normal human dermal fibroblasts (NHDF, PCS-201-012, ATCC, Manassas, VA) were grown in 10% fetal calf serum-supplemented DMEM. Cells were cultured in 6 and 12 well format and stimulated with rTWEAK (400 ng ml−1 murine rTWEAK for PAM212, 25-100 ng ml−1 human rTWEAK for human cells), 100 ng ml−1 species-specific rIL-13, 100 ng ml−1 species-specific rIL-17A, or combinations thereof (all from Peprotech, Rocky Hill, NJ). After 24 and 48 h, cells were harvested for real-time PCR with reverse transcription or flow cytometry analysis. To measure Fn14 surface expression, cells were stained with Fn14-APC (Miltenyi, Clone ITEM-4).
7
Culturing Primary Human Keratinocytes
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Primary human epidermal keratinocytes (normal human adult, HEKa) were obtained from American Type Culture Collection (PCS-200-011, ATCC) and cultured in dermal basal medium supplemented with a Keratinocyte growth kit (PCS-200-030 and PCS-200-040, also from ATCC). 293A cells were from Thermo Fisher and were maintained in Dulbecco’s Modified Eagle Medium (DMEM) (10-013-CMR, Corning) supplemented with 10% FBS (Gibco) and 1% penicillin-streptomycin solution (penicillin–Streptomycin 10,000 U/mL, Gibco). Cells were used at passage 2 to 3 post-thawing and routinely evaluated for contaminating mycobacterium using an in-house testing service.
8
Cultivation of Human Keratinocyte Cell Lines
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Normal human keratinocyte cell line human immortalized keratinocyte (HaCaT) and Normal Adult Human Primary Epidermal Keratinocytes (ATCC® PCS200011™) were purchased from CLS Cell Lines Service GmbH (Eppelheim, Germany) and from ATCC, respectively. HaCaT cells were maintained in RPMI-1640 medium (Sigma-Aldrich Chemie GmbH, Steinheim, Germany) containing 10 % (v/v) heat-inactivated fetal calf serum (FCS), 2 mM glutamine, 100 mg/mL streptomycin, and 100 U/mL penicillin. Normal Adult Human Primary Epidermal Keratinocytes were maintained in serum-free Dermal Cell Basal Medium (ATCC® PCS200030) supplemented with components of the keratinocyte growth kit (ATCC® PCS200040) that contains the following growth supplements: bovine pituitary extract (BPE), rhTGF-α, l -glutamine, hydrocortisone hemisuccinate, insulin, epinephrine, and apotransferrin. Both cell lines were grown at 37 °C in a 5 % CO2 humidified atmosphere. Cells in confluence were harvested, washed twice with phosphate-buffered saline (PBS), and detached from culture flasks by a brief treatment with 0.02 % EDTA solution (Sigma-Aldrich Chemie GmbH).
9
Keratinocyte Response to Inflammatory Stimuli
10
Culturing Human Melanoma Cell Lines
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Human melanoma cell lines HTB-72 (SKMEL-28) and CRL-1585 (C-32) were purchased from the American Type Culture Collection (ATCC, Manassas, VA, USA) and cryogenically preserved until use at passage numbers P20 or less for SKMEL-28, and P18 or less for C-32. The human melanoma cell line CRL-1619 (A-375) was obtained from the ATCC and was a gift provided by the Yaddanapudi laboratory. A-375 cells were cryogenically preserved until use at P16 or less. Human primary epidermal melanocytes (PCS-200-013) were purchased from the ATCC and cryogenically preserved until use at P6. Melanoma cells were cultured at 37 °C in 90% Dulbecco’s modified Eagle’s medium (DMEM) with 10% heat-inactivated fetal bovine serum (FBS) and 5% CO2. Primary melanocytes were cultured in primary melanocyte media composed of dermal basal medium (ATCC PCS-200-030), containing melanocyte growth factors (ATCC PCS-200-042), at 37 °C and 5% CO2.
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