I'm trying to count P. fluorescens using a hemocytometer (). Unfortunately they move too fast. Does someone know a unproblematic chemical to immobilize them? (Keep them in the hemocytometer for 2h and hoping that they run out of oxygen does not work...)
Additionally, I would like to differentiate between life and dead bacterias. I came across Trypan blue. Does somebody have any experiences colouring P. fluorescens adding Trypan blue? Or can you recommend another colouring agent that is less hazardous?
Thanks for your help!
Hi Fabienne
You can stain them with a regular gram-colouring, just avoid the de-inking step.
To slow them down, a highly viscous liquid like PEGs, Glycerol and Carbohydrate Thickeners, maybe with additional freezing might be suitable
Pseudomomnas are highly aerobic, I think 2 hours without oxygene will not be tolerated without basic damages, or error effects to MOs under optimum growth conditions.
Alternatively, you can us a flow-Cytometer or in your case, to distinguish between active and inactive cells or: make a simple and quick OD-Estimation and combine it with serial dilutions on Agar-Plates to estimate your living cells.....
Good Luck
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