Microvascular Endothelial Cell Growth Kit-BBE (ATCC® PCS-110-040)

Biosafety Level 1
Product Format frozen 1 kit
Storage Conditions -20°C (or -70°C for long-term storage)
Comments
The Microvascular Endothelial Cell Growth Kit-BBE contains components that when added to Vascular Cell Basal Medium (ATCC PCS-100-030) create a complete cell culture environment for microvascular endothelial cells isolated from normal human tissues. The final concentration for each component in complete endothelial growth medium is as follows:
  • Bovine brain extract (BBE): 0.2%
  • rh EGF: 5 ng/mL
  • L-glutamine: 10 mM
  • Heparin sulfate: 0.75 Units/mL
  • Hydrocortisone: 1 µg/mL
  • Ascorbic acid: 50 µg/mL
  • Fetal bovine serum: 5% 
Complete Growth Medium
  1. Obtain one growth kit from the freezer; make sure that the caps of all components are tight. 
  2. Thaw the components of the growth kit just prior to adding them to the basal medium. It is necessary to warm the L-glutamine component in a 37°C water bath and shake to dissolve any precipitates prior to adding to the basal medium. 
  3. Obtain one bottle of Vascular Cell Basal Medium (475 mL) 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 volume of each growth kit component, as indicated in Table 1 or 2, to the bottle of basal medium using a separate sterile pipette for each transfer. 
  6. Table 1. If using the Microvascular Endothelial Cell Growth Kit-BBE (ATCC® PCS-110-040), add the indicated volume for each component: 

    Component

    Volume

    Final Concentration

    Bovine Brain Extract (BBE)

    1.0 mL

    0.2%

    rh EGF

    0.5 mL

    5 ng/mL

    L-glutamine

    25.0 mL

    10 mM

    Heparin sulfate

    0.5 mL

    0.75 Units/mL

    Hydrocortisone hemisuccinate

    0.5 mL

    1 µg/mL

    Fetal Bovine Serum

    25.0 mL

    5%

    Ascorbic acid

    0.5 mL

    50 µg/mL

     

           Table 2. If using the Microvascular Endothelial Cell Growth Kit-VEGF (ATCC® PCS-110-041), add the indicated volume for each component:

    Component

    Volume

    Final Concentration

    rh VEGF

    0.5 mL

    5 ng/mL

    rh EGF

    0.5 mL

    5 ng/mL

    rh FGF basic

    0.5 mL

    5 ng/mL

    rh IGF-1

    0.5 mL

    15 ng/mL

    L-glutamine

    25.0 mL

    10 mM

    Heparin sulfate

    0.5 mL

    0.75 Units/mL

    Hydrocortisone hemisuccinate

    0.5 mL

    1 µg/mL

    Fetal Bovine Serum

    25.0 mL

    5%

    Ascorbic acid

    0.5 mL

    50 µg/mL

    Antimicrobials and phenol red are not required for proliferation but may be added if desired. The recommended volume of each optional component to be added to the complete growth media is summarized in Table 3.


    Table 3. 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

  7. 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.
  8.  Complete growth media should be stored in the dark at 2°C to 8°C (do not freeze). When stored under these conditions, complete media is stable for 30 days.
Volume 1 kit
Sterility Tests
Negative for bacteria, fungi and yeast
Functional Tests Rate of proliferation and morphology.
A Certificate of Analysis (COA) is available upon request for each lot of Microvascular Endothelial Cell Growth Kit.
C of A
Certificate of Analysis
Certificate of Analysis