I don't think it is a good idea to try to identify / isolate cancer stem cells in cell lines. Do you have access to primary leukemia samples or a leukemia mouse model?

Hi Daisy, as long as there are no established markers known, it will be very difficult to identify stem-cell like cells in these leukemia cell lines. c-KIT (CD117) or CD133 (for human cell lines) are other markers you could check. You could also try life cell-cycle analysis - maybe you can identify cells in G0 (somatic stem cells have usually a different cell cycle profile - however, this might not be true for cancer stem cells!). A very robust but also demanding approach would be to perform  limiting dilution transplantation assays of you leukemic cell lines into mice including serial-transplantation experiments - this way you can calculate what the frequency of leukemia initiating cells (= leukemic stem cells) is and with the serial transplantation you can test their self-renewal potential - this is very demanding and it also does not help to isolate a potential stem cell fraction!

Good luck!

Hi Daisy, I usually stain cancer cells with PKH26 fluorescent dye to evaluate cell proliferation and cell division. One of the most common uses of PKH26 is proliferation analysis based on dye dilution, therefore PKH26 dye is used for identification of quiescent or slowly dividing stem or progenitor cells in tissues. You can stain the leukemia cells with PKH26 dye and track its fluorescent dye using FCS, and then sort those non-dividing or slow-dividing cells.

Hope it is helpful for you.   

 Hi Dr. Schürch, thanks for your answer.  

I do have access to primary leukemia samples. However, I am planning to use transfection for mechanism study in LSC, since primary leukemia cells are difficult to maintain ex vivo, let alone withstanding further complicated transfection procedures. Or perhaps I should give it a try.

Best.

Daisy

Hi Dr. Baumgartner, Thank you for your answer.

It's true that lacking established leukemia stem cell markers, isolating and identifying LSC, especially in cell lines, can be quite difficult. Recent publications using different markers can only claim their cells to have stem cell properties, but definitely not TRUE LSC. And I would love to use the limiting dilution transplantation assays had I got the time and money ;) It's the most convincing methods so far. The life cell-cycle analysis sound reasonable.  Thank you so much!

Best, Daisy. 

Hello Han, Thank you for your answer.

Haven't heard of this dye before, so I just googled it ;)  I think it's quite reasonable and worth trying based on functional selection. Thank you, I will certainly give it a try.

Best, Daisy.