The hPDLF isolated from 16-year-old male were purchased from Lonza Group, Ltd (Basel, Switzerland). The cells were cultured and expanded in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% fetal bovine serum (FBS), 100 U/ml penicillin, and 100 μg/ml streptomycin at 37°C in a humidified atmosphere containing 5% CO2. Cells from passage levels 4–5 were used in the present study. The cell culture medium was refreshed every 3 days.
Hpdlf
Manufactured by Lonza
Sourced in Switzerland, United States
The HPdLF is a laboratory equipment designed for cell culture applications. It serves as a high-performance, low-footprint incubator that provides a controlled environment for the growth and maintenance of cell lines. The core function of the HPdLF is to maintain optimal temperature, humidity, and atmospheric conditions for cell culture processes.
Automatically generated - may contain errors
Lab products found in correlation
Trypsin edta, by Thermo Fisher Scientific (16 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (13 mentions)
Rpmi 1640 medium, by Thermo Fisher Scientific (9 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (5 mentions)
Heat inactivated fetal bovine serum, by Thermo Fisher Scientific (4 mentions)
T175 culture flask, by Thermo Fisher Scientific (4 mentions)
L ascorbic acid, by Merck Group (3 mentions)
Streptomycin, by Merck Group (2 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (2 mentions)
Hpdlfs, by Thermo Fisher Scientific (2 mentions)
Streptomycin, by Thermo Fisher Scientific (2 mentions)
Protran, by Cytiva (1 mentions)
Penicillin streptomycin neomycin, by Merck Group (1 mentions)
L glutamine, by Thermo Fisher Scientific (1 mentions)
Accutase solution, by Merck Group (1 mentions)
Accutase, by Merck Group (1 mentions)
Penicillin g, by Merck Group (1 mentions)
Fetal bovine serum (fbs), by Merck Group (1 mentions)
Amphotericin b, by Merck Group (1 mentions)
Ascorbic acid, by Merck Group (1 mentions)
17 protocols using hpdlf
1
Expansion and Maintenance of hPDLF Cells
2
Periodontal Ligament Fibroblasts and THP1 Cell Co-culture
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Pooled human periodontal ligament fibroblasts from several donors (HPdLF, Lonza, Basel, Switzerland) were grown in culture medium consisting of Dulbecco’s modified Eagle medium (DMEM; Thermo Fisher Scientific, Carlsbad, CA, USA) containing 4.5 g/L glucose, 10% heat-inactivated fetal bovine serum (Thermo Fisher Scientific, Carlsbad, CA, USA), 100 U/mL penicillin, 100 µg/mL streptomycin and 50 mg/L L-ascorbic acid at 37 °C, 5 % CO2 and 95% humidity. Cells were regularly passaged with 0.05% Trypsin/EDTA (Thermo Fisher Scientific, Carlsbad, CA, USA) when 75% confluence was reached. HPdLF of passage four to eight were used for experimental setups. For RNA and protein expression analysis, 1 × 105 cells were seeded per well of a 6-well plate and cultured to 75% confluency prior further treatment. For THP1 cell adherence assay, 50 HPdLF per mm2 were seeded on coverslips in 24-well plates and cultured to 75% confluence.
THP1 monocytic cells (DMSZ, Braunschweig, Germany) were cultured in RPMI 1640 medium (Gibco) containing 10 % FBS, 100 U/mL penicillin and 100 µg/mL streptomycin at 37 °C, 5 % CO2 and 95 % humidity. They were passaged weekly and seeded at a density of 1 × 106 cells in 20 mL medium in T175 culture flask (Thermo Fisher Scientific, Carlsbad, CA, USA).
THP1 monocytic cells (DMSZ, Braunschweig, Germany) were cultured in RPMI 1640 medium (Gibco) containing 10 % FBS, 100 U/mL penicillin and 100 µg/mL streptomycin at 37 °C, 5 % CO2 and 95 % humidity. They were passaged weekly and seeded at a density of 1 × 106 cells in 20 mL medium in T175 culture flask (Thermo Fisher Scientific, Carlsbad, CA, USA).
3
Western Blot Analysis of Stem Cell Markers
4
Culturing Human Periodontal Ligament Fibroblasts and Osteoblasts
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Human periodontal ligament fibroblasts (HPdLF, Lonza) and human primary osteoblasts (HOB, PromoCell) were cultured in Dulbecco’s modified Eagle’s medium (DMEM; Invitrogen) containing 10% FBS (Life Technology), 1% L-glutamine (Thermo Fisher Scientific), 1% penicillin/streptomycin/neomycin (Sigma Aldrich), 1% L-ascorbic acid (Sigma Aldrich) and 20 µg/mL dexamethasone (Sigma Adlrich) at 37 °C, 5% CO2 and 95% humidity. The starvation medium contained reduced FBS concentration (1%). When reaching confluence, cells were passaged by application of Accutase® solution (Sigma Aldrich). For experiments, cells were used at passages four to six.
5
Culturing Human Periodontal Ligament Fibroblasts
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Human periodontal ligament fibroblasts (HPdLF, Lonza) were cultured in Dulbecco’s modified Eagle’s medium (DMEM; Invitrogen) containing 4.5 g/L glucose, 1% L-glutamine, 10% FBS (Gibco), 1% penicillin/streptomycin and 1% L-ascorbic acid at 37 °C, 5% CO2 and 95% humidity. All cells were passaged regularly using either 0.05% Trypsin/EDTA (Invitrogen) or Accutase (Merck Millipore) and used for experiments at passages four to eight.
6
Evaluating Endodontic Sealers' Biocompatibility
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For examining the biological effects of the endodontic sealers, human periodontal ligament fibroblasts (hPDLF, Lonza, Basel, Switzerland), human osteoblasts (hOB, Promocell, Heidelberg, Germany), and human mesenchymal stem cells of male Caucasian origin (hMSC, Lonza) were used. hPDLF and hMSC were cultured in high glucose Dulbecco’s modified eagle’s medium (DMEM, gibco/life technologies), and osteoblast growth medium (OGM, PromoCell GmbH, Heidelberg, Germany) was used for the cultivation of hOB. All media were supplemented with 10% fetal bovine serum (FBS, Merck, Darmstadt, Germany), 100 U/ml of penicillin G and 100 µg/ml streptomycin (Merck). All experiments were performed with cells at passage 4–8 and all cell types were incubated at 37 °C in a humidified atmosphere containing 5% CO2.
7
Osteogenic Differentiation of Human PDL Fibroblasts
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The human periodontal ligament fibroblast (hPDLF) was purchased from Lonza (Lonza, Basel, Switzerland). The cells were resuspended in culture medium (DMEM; Gibco, Carlsbad, CA, USA) supplemented with 10% fetal bovine serum (Gibco), 1% penicillin/streptomycin (Gibco), and 0.025 μg/mL amphotericin B (Sigma–Aldrich, St. Louis, MO, USA). The cells were cultured in a humidified 5% CO2 incubator at 37 °C. When the hPDLF reached 85% confluence, 50 μg/mL ascorbic acid (Sigma–Aldrich), 10 mM β-glycerophosphate (Sigma–Aldrich), and 100 nM dexamethasone (Sigma–Aldrich) were added in the culture medium to induce osteogenic differentiation.
8
Periodontal Ligament Fibroblast and Monocyte Cell Culture
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Commercially acquired human periodontal ligament fibroblast (HPdLF, Lonza, Basel, Switzerland) were grown in culture medium consisting of Dulbecco’s modified Eagle medium (DMEM; Thermo Fisher Scientific, Carlsbad, CA, USA) containing 4.5 g/L glucose, 10% heat-inactivated fetal bovine serum (Thermo Fisher Scientific, Carlsbad, CA, USA), 100 U/mL penicillin, 100 µg/mL streptomycin, and 50 µg/mL L-ascorbic acid at 37 °C, 5% CO2 and 95% humidity. Cells were passaged when reaching a confluency of 75% with 0.05% Trypsin/EDTA (Thermo Fisher Scientific, Carlsbad, CA, USA). For experiments, HPdLF of passage four to eight were used.
THP1 monocytic cells (DMSZ, Braunschweig, Germany) were cultured in RPMI 1640 medium (Thermo Fisher Scientific, Carlsbad, CA, USA) containing 10% FBS, 100 U/mL penicillin, and 100 µg/mL streptomycin at 37 °C, 5% CO2, and 95% humidity. The non-adherent cells were passaged regularly after seven days and seeded at a density of 1 × 106 cells in 20 mL medium in T175 culture flask (Thermo Fisher Scientific, Carlsbad, CA, USA). For this, cells were pelleted by centrifugation for 5 min at 1000× g and diluted in 1 mL RPMI culture medium prior to cell counting in a hemocytometer (Neubauer Chamber Improved, Avantor, Radnor, PA, USA).
THP1 monocytic cells (DMSZ, Braunschweig, Germany) were cultured in RPMI 1640 medium (Thermo Fisher Scientific, Carlsbad, CA, USA) containing 10% FBS, 100 U/mL penicillin, and 100 µg/mL streptomycin at 37 °C, 5% CO2, and 95% humidity. The non-adherent cells were passaged regularly after seven days and seeded at a density of 1 × 106 cells in 20 mL medium in T175 culture flask (Thermo Fisher Scientific, Carlsbad, CA, USA). For this, cells were pelleted by centrifugation for 5 min at 1000× g and diluted in 1 mL RPMI culture medium prior to cell counting in a hemocytometer (Neubauer Chamber Improved, Avantor, Radnor, PA, USA).
9
Overexpression of BCOR Mutations in Fibroblasts
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A wild-type (WT) BCOR plasmid, mutated (Mut) BCOR plasmid with a c.3668delC frameshift mutation in the BCOR gene, and an empty plasmid described in our previous study were used (Surapornsawasd et al., 2015 (link)). Primary human PDL fibroblasts, HPdLF (Lonza, Walkersville, MD, United States), and COS-7 cells were grown and cultured in SCGM BulletKit™ medium (Lonza) and α-MEM (Wako), respectively; they were kept in a humidified 5% CO2 atmosphere at 37°C. Overexpression of each plasmid was performed using Lipofectamine P3000 reagent (Invitrogen, Van Allen Way Carlsbad, CA, United States) according to the manufacturer’s instructions. After 24 h, immunofluorescence and cell proliferation assays were performed. All experiments using the expression plasmids were approved by the Recombinant Experiment Committee of our institution (G2020-021A).
10
Porphyromonas gingivalis LPS Induced Inflammation in Human PDL Fibroblasts
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Human PDL fibroblasts (HPDLFs) were obtained from Lonza and were cultured with SingleQuots Supplements (CC‐4181, Insulin, hFGF‐β, GA‐1000, and fetal bovine serum) at 37°C. HPDLFs were subcultured when they became 70%–80% confluent and cells at passages 8–9 were used for these experiments. The cells were incubated in the presence or absence of 1 µg/ml Porphyromonas gingivalis LPS (LPS‐PG Ultrapure; InvivoGen) to induce an inflammatory response for 24 h.
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