IMR-90 Cell culture protocol (Gabrielle): adapted from the ATCC website


            MEM-alpha + Glutamax (500mL) with:

  1. 10% FBS (50mL)
  2. Pen/strep (5mL)


Day 1:

  1. Cells are kept in the orange Liquid Nitrogen tank, Rack 3, Box 3 (counting from bottom up)
  2. Do NOT warm media in a water bath; media should be in room temp as to not stress the cells
  3. Thaw the vial in a water bath
    1. Do NOT let the vial completely thaw in the water bath
    2. Keep lid out of water bath
    3. Gently move vial side to side as thawing
    4. It should be less than 1-2min
  4. Spray tube with EtOH to decontaminate, bring into hood
  5. Slowly add RT media to the cells
    1. For 1mL frozen cells use 9mL of fresh media
  6. Transfer thawed cells into the tube of fresh media as they become fully thawed
  7. Transfer 10mL of media+cells into a 10cm dish
  8. Label lid with passage, cell type, date, and initials
  9. Place dish with cells in the incubator
    1. 37C, 95% Air, 5% CO2

Day 2:

  1. Check on cells by using the microscope
  2. Change media to remove any remaining cryoprotective agent
  3. Remove old media
  4. Add 10mL of warmed media to the plate gently to not lift any live cells

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