I am currently staining and fixing a range of bacteria for a glycobiologist.
I am fixing the bacteria in 4% PFA and then washing with PBS. After fixing Lactobacillus spp. in PFA, it wont pellet. I have centrifuged it at 10000g for 5 minutes.
Does anyone perhaps know why this is? What fixation method should I use instead.
David Ian Leavesley
PFA is a crosslinking reagent! You have cross-linked your bacteria forming a networked gel/sponge. They will not sediment at 10,000 g in 5b minutes!
Suggest that you use cold methanol, or similar, to fix your bugs.
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