problem with cell cycle analysis
Posted 28 June 2010 - 08:50 PM
I am novice to cell cycle anaysis and done this procedure ( Propidium iodide staining) for the first time in A431 cancer cell lines.
my results were discouraging
the person who analysed my samples for cell cycle analysis told me that the cells are beyond their duplicate forms . what does it mean? (multinucleated cells?)
also she had to lower the voltage to analyse histograms. what it suggests?
Please help me to solve my doubt and how can I overcome these problems?
method I followed-
- seed 10*5 to 10*6 cell in 50 to 80mm petri dish
-after overnight incubation I added drug to petri dishes
after 48 hrs I fixed the cell in ethanol(stored at -20 deg celcius)
- kept cells in 4 deg celcius.
next day did PI staining.
Posted 28 June 2010 - 10:48 PM
the cells u r using hav the property of forming clumps very quickly. So when u r acquisitioning the sample by flow.. vortex properly before.. and in between also... maximum u can use 1 million cells for staining.. and after fixation with ethanol .. store at -20c for overnight to 48 hrs will help in better results...
Posted 01 July 2010 - 06:53 AM