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Cynomolgus Monkey Coronary Artery Endothelial Cells

Cynomolgus Monkey Coronary Artery Endothelial Cells
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Price: $0.00
Cat. #: MK-6093

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Cynomolgus Monkey Coronary Artery Endothelial Cells

Catalog No. MK-6093

Suggested Medium: Catalog No. MK1168  Endothelial Cell Medium /w Kit

 

Product Description

Cynomolgus Monkey Coronary Artery Endothelial Cells from Cell Biologics are isolated from tissue of Cynomolgus Monkey. Monkey Coronary Artery Endothelial Cells are grown in T25 tissue culture flasks pre-coated with gelatin-based solution for 0.5 hour and incubated in Cell Biologics’ Culture Complete Growth Medium generally for 3-7 days. Cultures are then expanded. Prior to shipping, cells are detached from flasks and immediately cryo-preserved in vials. Each vial contains at least 1x106 cells per ml and are delivered frozen. The method we use to isolate primary endothelial cells was developed based on a combination of established and our proprietary methods. These cells are pre-coated with PECAM-1 antibody, following the application of Dynabeads pre-coated with secondary antibody. 

Product Testing

Monkey Coronary Artery Endothelial Cells are tested for uptake of Dil-Ac-LDL, a functional marker for endothelial cells. Monkey Coronary Artery Endothelial Cells are negative for bacteria, yeast, fungi and mycoplasma, and can be expanded for 3-6 passages at a split ratio of 1:2 under the cell culture conditions specified by Cell Biologics. Repeated freezing and thawing of cells is not recommended.

Laboratory Applications

Monkey Coronary Artery Endothelial Cells can be used in assays of cell to cell adhesion, migration, vascular tube formation. Standard biochemical procedures performed with endothelial cell cultures include RT-PCR, Western blotting, immunoprecipitation, immunofluorescent staining or immunofluorescent flow cytometry or generating cell derivatives for desired research applications.

Storage of Cell Biologics’ Products

Cell Biologics ships frozen cells on dry ice. On receipt, immediately transfer frozen cells to liquid nitrogen (-180 °C) until ready for experimental use.  Live cell shipment is also available on request.

Never can primary cells be kept at -20 °C.

Authorized Uses of Cell Biologics’ Products

Monkey Coronary Artery Endothelial Cells from Cell Biologics are distributed for research purposes only. Our products are not authorized for human use, for in vitro diagnostic procedures or for therapeutic procedures.  Transfer or resale of any Cell Biologics’ cells or products from the purchaser to other markets, organizations or individuals is prohibited by Cell Biologicswithout the company’s written consent.  Cell Biologics’ Terms and Conditions must be accepted before submitting an order.

Disclaimer

Investigators should handle the cells that they receive from Cell Biologics with caution and treat all animal cells as potential pathogens, since no test procedure can completely guarantee the absence of infectious agents.

Warranty and Liability

CELL BIOLOGICS’ guarantee applies only to your purchase of CELL BIOLOGICS' cells with CELL BIOLOGICS’ Media and Coating Solution for appropriate cell culture and cell testing, following CELL BIOLOGICS’ online protocols within 35 days from the date of product delivery.


Primary Cell Culture Protocol

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All cell culture procedures must be conducted in a bio-safety cabinet.

Any and all media, supplements, and reagents must be sterilized by filtration through a 0.2 µm filter.

Use aseptic technique to prevent microbial contamination.

Cryo-preserved cells must be stored in liquid nitrogen or seeded immediately upon arrival.

Medium:

Review the information provided on the Cell Biologics website about appropriate culture media (e.g. serum and other supplements).  Use pre-warmed (37°C) cell culture media (30-50 ML) to recover cryo-preserved cells and when changing media or splitting cells.

Coating of flasks or dishes:

Coat sterile culture dishes or flasks with Gelatin-Based Coating Solution (Cell Biologics, Catalog No. 6950) for 10-30 min and then aspirate the excess solution before seeding cells.

Cell recovery from cryovial:

  • Quickly thaw cells in cryo-vial by incubating them in a 37°C water bath for <1 min until there is just a small bit of ice left in the vial.
  • Promptly remove the vial and wipe it down with 70% ethanol.
  • Transfer cells from the vial to a sterile centrifuge tube. Add 8-10 ml of pre-warmed Cell Biologics Cell Culture Medium. Flush the vial with an additional 0.5-1 ml of medium to ensure complete transfer of cells to the centrifuge tube.
  • Centrifuge cells at 200 g for 5 minutes.
  • Aspirate the supernatant and resuspend the cell pellet in 6 ml of Cell Biologics’ Cell Culture Growth Medium.
  • Add resuspended cells into a T25 flask pre-coated with Gelatin-Based Coating Solution (Cell Biologics, Catalog No. 6950).
  • Place the T25 flask in a humidified, 5%-CO2 incubator at 37°C. Change medium 3-6 hours after thawing cells to ensure >90% cells are attached. 
  • Also change culture media the following day to remove non-adherent cells and replenish nutrients.
  • Change cell culture medium everyday when cells are >70% confluent.
  • Cells should be checked daily under a microscope to verify appropriate cell morphology.

Expansion of cultured mouse primary cells:

  • Flush the adherent layer with a 5 ml sterile pipette 3 times to dislodge loosely attached cells.
  • Remove and discard the cell culture media from the flask.
  • Wash adherent cells 2 times with 10 ml of sterile PBS (1X) without calcium and magnesium to remove nonadherent cells or fraction.
  • Remove and discard the wash solution from the flask.
  • Incubate cells with warm (37°C) 0.05% Trypsin-EDTA (1X) solution (Invitrogen catalog number 25300) for 1 minute. Use 2.0 ml of 0.05% Trypsin-EDTA solution when collecting cells from T75 flasks, and 1.0 ml when using T25 flasks. As soon as cells have detached (the flask may require a few firm gentle taps), add 8-10 ml of Cell Biologics Cell Culture Medium supplemented with 5-10 % FBS to a T25 or T75 flask (the FBS will neutralize the trypsin).
  • Plate cells in fresh flasks or plates precoated with Gelatin-Based Coating Solution in a humidified, 5%-CO2 incubator at 37°C. Change medium 3-6 hours after seeding cells to ensure >90% cells are attached. 
  • Also change culture media the following day to remove non-adherent cells and replenish nutrients.
  • Cells should be checked daily under a microscopy to verify appropriate cell morphology.
  • Change culture medium every 24-48 hours. The medium should be changed every day when cells are >70% confluent to remove non-adherent cells and replenish nutrients.
  • Pre-wash cells with 1X PBS 2 times whenever replacing the medium.
  • We recommend splitting primary cells at the follow ratio:
  • The recommended split ratio for primary murine cells is 1:2.
  • A confluent monolayer of primary cells grown in a T75 flask may be expanded on one or two 6-well plate ready for use in experiments under the cell culture conditions specified by Cell Biologics.

Procedure for Freezing Cells

Materials:

  • 1X Phosphate Buffered Saline (PBS-1X)
  • 0.05% Trypsin-EDTA (1X) solution (Invitrogen catalog number 25300)
  • Tissue Culture Media
  • Cold Freezing Media (10% DMSO, 20% FBS and 70% culture medium)
  • Labeled Cryovials
  • Confluent cells

         ---------------------------------------

  • Flush the adherent cell layer with a 5 ml sterile pipette 3-5 times to dislodge loosely attached cells.
  • Remove and discard the cell culture media from the flask.
  • Wash adherent cells 2-3 times with 10 ml of sterile PBS (1X) without calcium and magnesium to remove nonadherent cells or fraction.
  • Remove and discard the wash solution from the flask.
  • Incubate cells with warm (37°C) Trypsin-EDTA (1X) solution for 1 minute. Use 2.0-3.0 ml of 0.05% Trypsin-EDTA solution when collecting cells from T75 flasks, and 1.0-1.5 ml when using T25 flasks. As soon as cells have detached (the flask may require a few firm gentle taps), add 10 ml of Cell Culture Medium supplemented with 5-10 % FBS to the flask (the FBS will neutralize the trypsin).
  • Centrifuge the cell suspension at 200 g for 5 minutes.
  • Remove supernatant with sterile Pasteur pipette.
  • Quickly re-suspend pellet by adding 1.0 ml freezing media per vial to be frozen.
  • Place Cryovials in Nalgene "Mr. Frosty" freezing container containing100% isopropyl alcohol at -70-80 °C for 24 h.
  • Transfer vials to liquid N2 tank for indefinite storage.

We recommend freezing primary cells at the follow ratio:

  • A confluent primary endothelial cells grown in a T75 flask may be frozen in 3 cryovials.
  • A confluent primary endothelial cells grown in a T25 flask may be frozen in 1 cryovial.
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