What is the best choise for unstick differentiated THP-1 cells? We tried EDTA, but I would like to observe some markers of differentiated by flow cytometry. Unfortunatelly, after EDTA THP1cells have circle shape again and I think that EDTA change some surface properties. Do you have any experience?
Hi Veronika,
I completely agree with Lukas Pawig. I used to scrap the activated Thp1 cells feeded with labeled bacteria for FACS analysis. Whenever surface properties needs to be studied by flow cytometry, scraping is a better method than trypsinizing.
Hope it will help you
Biswaranjan
Hi Veronika,
as stupid as it sounds, but I was successful in my experiments with gently scraping off the cells with a sterile cell scraper (e.g. by Corning). As you said, Trypsin or EDTA Treatment is too harsh on the cells, since they are really really sticky. You will of course loose some cells with scraping, but you can easily gate the defect cells out in flow cytometry. I evaluated CD68 as a marker for macrophages and PMA-differentiated THP1 had significantly higher levels than untreated THP-1.
You could at least give this a try, it's a very simple method.
Hi Veronika,
I completely agree with Lukas Pawig. I used to scrap the activated Thp1 cells feeded with labeled bacteria for FACS analysis. Whenever surface properties needs to be studied by flow cytometry, scraping is a better method than trypsinizing.
Hope it will help you
Biswaranjan
Hi Veronika,
Since it is known trypsin affects the expression of various cell surface markers, in our lab we use Accutase to detach adherent cells. Accutase is much gentler but very efficient in detaching cells. In comparison to trypsin, it works better for follow up flow cytometry analysis.
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