I'm working on THP-1 monocytes. I thawed them last week to start my culture, but I can't count them. I count in Mallassez cells with trypan blue. I can see by microscopy that my cell density is increasing, but I can't see anything in Malassez cells when I try to count them....
Do you have any advice on how to count these cells?
Hello Chloé Chevalier ,
This might be a silly answer, but depending on the volume of cells you are working with, if your suspension is not well homogeneized, you might get only a few cells in your sample, and when you dilute it with your trypan blue solution, you "loose" them on your Malassez cell.
Also, compared to other cells in suspension, THP-1 are small cells that regularly aggregate in quite large clusters when they reach a certain density, so maybe your density is not enough to be able to see something ? Do you observe any clusters, or only isolated cells ?
And on your Malassez, do you see absolutely nothing, or only a small number of cells ?
I don't know if you have the possibility to use an automatic cell counting device ? I know this is not always the case in some labs, but it really is a game-changer ! If you can't, maybe you have the possibility to try and use another manual cell counting method (with the appropriate cell of course), like Burker, just to see if you see something or not.
I don't remember the specifics of Malassez cell counting because I have the luxury of a ViCell counting device at my facility, but maybe you could try to dilute your sample a bit less ?
Hope this helps !
You can pellet down the suspension and dilute with a lesser volume of media. Supposing you have 20 ml of suspension culture. Then, after centrifugation and discarding the previous media, you can add 5 ml of media, then take cells and trypan blue to count with the cell counter.
I hope this will help you.
1. Try to dilute your sample.
2. Viable THP-1 cells have a shiny and glowing appearance; they may be distinguished from dead cells without the need for trypan blue staining.
I hope this will help you.
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