Apoptosis assay - Trypsin V Scraping (Feb/21/2007 )

Hello All

I was wondering wether trypsinising or scraping cells is better for harvesting cells to analyse apoptosis (using Annexin and Propidium iodide staining). Any advice?

Many thanks in advance

-oaktree-

QUOTE (oaktree @ Feb 21 2007, 11:16 AM)

Hello All

I was wondering wether trypsinising or scraping cells is better for harvesting cells to analyse apoptosis (using Annexin and Propidium iodide staining). Any advice?

Many thanks in advance

why detaching cells and not directly measuring annexin V and PI labeling in fluorescence microscopy or fluorescence plate reader (for cell culture in 98 or 384-wells)?

-The Bearer-

QUOTE (The Bearer @ Feb 21 2007, 10:18 AM)

QUOTE (oaktree @ Feb 21 2007, 11:16 AM)

Hello All

I was wondering wether trypsinising or scraping cells is better for harvesting cells to analyse apoptosis (using Annexin and Propidium iodide staining). Any advice?

Many thanks in advance

why detaching cells and not directly measuring annexin V and PI labeling in fluorescence microscopy or fluorescence plate reader (for cell culture in 98 or 384-wells)?

I am measuring cell death using FACS to analyse early and late stage apoptosis by combining propidium iodide and annexin staining, therefore I need to detach the cells but I am concerned about the effect trypsin or scraping may have on the cells and if this could affect the result.

-oaktree-

hi,

you may try citric saline for recovery of adherent cells
http://www.protocol-online.org/forums/inde...6&hl=citric

I have tried DiOC6(3) staining after citric saline and both apoptotic and viable cells were clearly distinguishable, in contrast to trypsin and scraping methods...

Seb

-tryptofan-

QUOTE (tryptofan @ Feb 22 2007, 09:24 AM)

Thanks a million, I'll give it a go!

-oaktree-