Cell lysis with proteinase k - (Jun/18/2012 )
I'm looking at GAG quantification from cell cultures using either DMMB or Alcian blue however I really need to normalise across different cell lines so would like to normalise to DNA content. Would a proteinase K digest (in 100mM ammonium acetate) followed by freeze thaw (1 or more cycles) be sufficient to release the cellular DNA so I can split the sample for quantification by picogreen without further processing and for GAG analysis? I don't really want to add other lysis agents as they may interfere with the GAG analysis.
You can normalize baased on protein amount. Do protein assay and start with same amount of proteins for different cell lines.
Freeze-thaw alone is usually sufficient to disrupt the nuclear membrane.