Asynchronous Population – cells all throughout the cell cycle

Synchronous Population – group of cells that are in the same part of the cell cycle

Tests :

Synchronzining Cells so we know where they are in the Cell Cycle:

Mitotic Harvest:

Petri Dish : contains a growth medium and is placed in a temperature condusive to growth (above room temperature)

• Works on the premise that cells have a natural cohesivness that will make them stick to the bottom of the Petri Dish
• During Mitosis cells do not stick together as much therefore by lightly tapping the Petri Dish you will loosen all of the cells that are undergoing Mitosis
• Then these cells are transferred to another Petri Dish (this is syncronization of cells)
• We can now study the cells in both Petri Dishes each at different parts of the cells cycle
• Note: not all of the cells will transfer from dish to dish, some will die due to this process  (Example : 70 out of 100 cells survive,  the 70 that survive are called the Platting Factor and will be the new value for 100% survival)

This process is very primitive so new ways were found to collect cells

Irradiate Cells (G2M block):

Cells are irradiated with 60 - 300 rads. This will delay all cells going from G2 – Mitosis (synchronizer)

• Radiation at therapeutic doses create a block for cells going from G2 to Mitosis (this is called the G2M Block)
• G2M Block – mitotic delay due to a dose of radiation (dose in the therapeutic range 60-300 cGy) also the cells that are in Mitosis will die

1. Mitotic Cells Die
2. G2 Cells stop progressing because of the G2M Block
3. All other cycles in other cycles continue to progress through the cells cycle
4. When they reach G2 they are all synchronized until the delay ends and all of the cells will progress together through G2 (they are now synchronized)

This delay will end in 5 hours (this is the premise for B.I.D. treatments, so after the 5 hour wait all cells will be going through Mitosis and this is when the second treatment of radiation is given)

• Must wait 5-6 hours in between treatments
• Used mostly for head and neck patients
• Only works at therapuitic doses

 

Chemicals Used:

Hydroxyurea : puts a G1 block on the cell cycle (synchronizer)

• after the cells are blocked from going through G1 Phase they are now synchronized and can be tracked
• Hydroxyurea is very toxic and will attack cells that are already in S Phase
• It is an anti-metabolite that was used as a chemotherapy drug

Labeling Cells so we can see where they are at the Cell Cycle:

H³Tdr – Tritiated Thymidine (is a labeler) : it is  a radioactive neuclotide

• during the S Phase (Synthesis) the cells will pick up the Tritiated Thymidine instead of thymine
• then these cells are placed in  a dark room and an emulsion is put over it and is then filmed (process takes about 30 days)
• cells can are tracked because the Tritited Thymadinine will show on the films at what part of the cell cycle the cells is in

BUDR – bromodeoxyuridine  (is a labeler): is another radioactive labeler like tritiated thymadine but instead of replacing thymine it will replace uracil

• It works faster than the tritiated thymadine
• Also makes hypoxic cells