How do you thaw HepG2 cells?
Thawing and Plating HepG2 Cells Remove the vial of cells from dry ice or storage unit. Defrost the vial of cells in a 37oC water bath with constant, moderate agitation, until ice in the ampoule is no longer visible. 2. Continue to warm the ampoule in the water bath for 30 seconds with gentle agitation.
Where are HepG2 cells?
HepG2 is an immortalized cell line consisting of human liver carcinoma cells, derived from the liver tissue of a 15-year-old Caucasian male who had a well-differentiated hepatocellular carcinoma, which is the fifth most-common cancer worldwide.
How do you revive a frozen cell line?
Guidelines for thawing cells
- Thaw frozen cells rapidly (< 1 minute) in a 37°C water bath.
- Dilute the thawed cells slowly before you incubate them, using pre-warmed growth medium.
- Plate thawed cells at high density to optimize recovery.
- Always use proper aseptic technique and work in a laminar flow hood.
Why must thawing of cells be done quickly?
Cell freezing needs to occur at a slow, controlled cooling rate. In contrast, cell thawing works best when it is done quickly because the disappearance of ice around the cell does not have the same damaging effects as ice formation during cryopreservation.
How fast do hepg2 cells grow?
Cells were found to clump extensively. Care were taken to break the clumps by vigorous pipetting during passages, yet the clumps were developed by the following day. Growth rate was also found to be low with population doubling time of 10 days.
How many cells should you freeze down?
Using a pipette, remove the supernate down to the smallest volume without disturbing the cells. Resuspend cells in freezing medium to a concentration of 1 x 107 to 5 x 107 cells/mL for serum-containing medium, or 0.5 x 107 to 1 x 107 cells/mL for serum-free medium. Aliquot into cryogenic storage vials.
Can you use cells immediately after thawing?
I would recommend using them after at least 2 passage after thawing, so that they are fully recoverred from the freeze/thawing stress. I agree with Kevin, it’s not advisable to use them until they have been passaged at least twice, since they need to recover and resume their normal cell cycle.
What is the difference between Emem and DMEM?
The main difference between DMEM and EMEM is that DMEM contains four times more vitamins and amino acids and two to four times more in comparison to the EMEM formula whereas EMEM is based on six salts and glucose. Furthermore, DMEM contains iron in the form of ferric sulfate.
Can you culture cells without CO2?
Mammalian cells in bicarbonate-based media require 5% CO2 to maintain physiological pH. Without a CO2 supply, cells are adversely affected within five minutes. To minimize pH change, you can supplement media with 25 mM HEPES buffer when possible.
What is the pathophysiology of HepG2?
HepG2 is an immortalized cell line consisting of human liver carcinoma cells, derived from the liver tissue of a 15-year-old Caucasian male who had a well-differentiated hepatocellular carcinoma. Hepatocellular carcinoma is the fifth most-common cancer worldwide. The morphology of HepG2 cells is epithelial and contains 55 chromosome pairs.
What is the protocol for transfection of HepG2 cells?
Transfection Protocol An optimized protocol to transfect HepG2 cells in a 24-well plate is described below: Plate 7,500-12,000 HepG2 cells per well in 0.5 ml of complete growth medium 12-24 hours prior to transfection Wash with 1xPBS and add 0.5 ml of fresh growth medium
What is the best way to propagate HepG2?
Cell Culture SOP: Propagation of HepG2 2 5) Immediately remove cells and pellet at 500 xg for 5 minutes (4oC). 6) Wash cells 2X with 1X PBS. 7) Gently re-suspend cell pellet in warm medium.
What is the origin of the HepG2 cell line?
HepG2 Cell Line Origin and Characteristics. HepG2 is an immortalized cell line consisting of human liver carcinoma cells, derived from the liver tissue of a 15-year-old Caucasian male who had a well-differentiated hepatocellular carcinoma, which is the fifth most-common cancer worldwide. The HepG2 cell line is commonly used in drug metabolism