Primary Gingival Fibroblast; Normal, Human, Adult (HGF) (ATCC® PCS-201-018)

Organism: Homo sapiens, human  /  Tissue: gingival  /  Cell Type: fibroblast

Organism Homo sapiens, human
Tissue gingival
Cell Type fibroblast
Morphology spindle-shaped; cells are bipolar and refractile
Growth Properties adherent
Biosafety Level 1

Biosafety classification is based on U.S. Public Health Service Guidelines, it is the responsibility of the customer to ensure that their facilities comply with biosafety regulations for their own country.

Age 60 years
Gender female
Ethnicity Caucasian
Applications

Human gingival fibroblasts (hGF) could potentially be an alternative source of mesenchymal stem cells (MSC) for regenerative medicine studies as they share similar morphology, CD markers, and differentiation lineage.

Product Format frozen
Storage Conditions liquid nitrogen vapor phase
Images ATCC PCS-200-014 FB Marker Images Cell Micrograph of ATCC PCS-201-018 cells
Comments Cryopreserved human oral fibroblast derived from adult gingival tissue. Oral fibroblasts are cryopreserved in P2: After isolation from tissue the cells are cultured for two passages and then cryopreserved. Vials contain 500,00 cells in one mL cryopreservation solution.
Complete Growth Medium
  1. Obtain one vial of Primary Normal Gingival Fibroblast Cells (ATCC PCS-201-018) from the freezer; make sure that the caps of all components are tight.
  2. Thaw the components of the growth kit (ATCC PCS-201-041) just prior to adding them to the basal medium (ATCC PCS-201-030).
  3. Obtain one bottle of Fibroblast Basal Medium (485 mL; ATCC PCS-201-030) from cold storage.
  4. Decontaminate the external surfaces of all growth kit component vials and the basal medium bottle by spraying them with 70% ethanol.
  5. Using aseptic technique and working in a laminar flow hood or biosafety cabinet, transfer the indicated volume of each growth kit component, as indicated in Table 1, to the bottle of basal medium using a separate sterile pipette for each transfer.

 Table 1. If using the Fibroblast Growth Kit-Low Serum, add the indicated volume for each of the following components

    Component

    Volume

    Final Concentration

    rh FGF b

    0.5 mL

    5 ng/mL

    L-glutamine

    18.75 mL

    7.5 mM

    Ascorbic acid

    0.5 mL

    50 µg/mL

    Hydrocortisone Hemisuccinate

    0.5 mL

    1 µg/mL

    rh Insulin

    0.5 mL

    5 µg/mL

    Fetal Bovine Serum

    10.0 mL

    2%

     

 

Antimicrobials and phenol red are not required for proliferation but may be added if desired. The recommended volume of either of the optional components (GA solution or PSA solution) to be added to the complete growth media is summarized in Table 2.

Table 2. Addition of Antimicrobials/Antimycotics and Phenol Red (Optional)

Component

Volume

Final Concentration

Gentamicin-Amphotericin B Solution

0.5 mL

Gentamicin: 10 µg/mL

Amphotericin B: 0.25 µg/mL

Penicillin-Streptomycin-Amphotericin B Solution

0.5 mL

Penicillin: 10 Units/mL

Streptomycin: 10 µg/mL

Amphotericin B: 25 ng/mL

Phenol Red

0.5 mL

33 µM


  1. Tightly cap the bottle of complete growth medium and swirl the contents gently to assure a homogeneous solution. Do not shake forcefully to avoid foaming. Label and date the bottle.
  2. Complete growth media should be stored in the dark at 2°C to 8°C (do not freeze). When stored under these conditions, complete growth media is stable for 30 days.
Culture Conditions
Atmosphere: air, 95%; carbon dioxide (CO2), 5%
Temperature: 37°C
Subculturing
Volumes used in this protocol are for 75 cm2 flask; proportionally reduce or increase amount of dissociation medium for culture vessels of other sizes. Corning® T-75 flasks (catalog #430641) are recommended for subculturing this product.
  1. Remove and discard culture medium.
  2. Rinse the cell layer with DPBS solution for 2 minutes to remove all traces of serum that contains trypsin inhibitor.
  3. Add 5.0 to 7.0 mL of Trypsin-EDTA solution to the flask and incubate at 37°C .Observe cells under an inverted microscope until cell layer is dispersed (usually within 4 to 6 minutes).
    Note: To avoid clumping do not agitate the cells by hitting or shaking the flask while waiting for the cells to detach. Add 5.0 to 7.0 mL of Trypsin Neutralizing Solution (ATCC® PCS-999-004™) Centrifuge at 125 x g; 10 ± 2 minutes. Discard supernatant and resuspend the cell pellet with 8 mL of complete growth media. Gently break cell pellet by pipetting repeatedly.
  4. Count cells. Seed 2,500 to 5,000 viable cells per cm2. Add appropriate volume of the cell suspension to new culture vessels.
  5. Incubate cultures at 37°C.
Change media every 2-3 days
Subculture when cells reach 75-80% confluence. Seeding density should be 2,500 to 5,000 viable cells per cm2
Sterility Tests Bacteria and Yeast: No growth
Mycoplasma: No growth
Viral Testing Hepatitis B: None detected
Hepatitis C: None detected
HIV-1: None detected
HIV-2: None detected
Viability ≥ 70%
Population Doubling Capacity 15 PDL
C of A
Certificate of Analysis
Certificate of Analysis