I have made many attempts to raise adherent macrophage-like HL-60 cells in order to run phagocytosis assays and flow, but I cannot seem to lift them without killing or damaging the vast majority of the cells. Does anyone have a trustworthy protocol for induction and resuspension that includes specific details?
Hi
Maybe you will find an answer to your question? Please see this article:
J Cell Physiol. 1989 May;139(2):313-9.
Effects of fluorescent derivatives of TPA on HL60 cells: dissociation between the differentiation-induced and protein kinase C activity.
Phuong, thank you for responding!
I see that your method for harvesting cells is very simple in that you grew them in a 100mm culture dish and resuspended with a rubber policeman in the presence of calcium and magnesium free PBS.
Were there any other reagents you used in the process or was it just mechanical lifting with the rubber policeman?
Did you use trypan blue staining to determine viability of the cells after resuspending them at 10^7 cells/ml? Are they viable?
Can you please send me a link to the rubber policeman you used? The one I am currently using may be significantly different.
ref for a rubber policemen :
United Scientific Supplies Stirring Rods with Rubber Policeman
Good luck
Thank you for your help! I was able to successfully raise the cells and maintain viability using the method you suggested, but I am having an issue with achieving adherence of a large percentage of my cell population. Please provide me with the cell density and media volume you used for the induction with PMA in order to achieve a cell density of 10^7/ml following resuspension.
- Badges
- Science topic
- Similar topics
- Biological Science
- Molecular Biology
- Molecular Biological Techniques