Hello Shahin,
one option would be to use the SYTO BC dye in combination with FACS. SYTO BC is a high-affinity nucleic acid stain that penetrates cells easily, producing a fluorescent signal.
There is a commercially available kit that might be suitable (it also contains labelled microspheres for calibration of the signal):
http://www.lifetechnologies.com/order/catalog/product/B7277
Although this is optimized for use with bacteria it might also be applicable to protozoans.
Good luck with your experiments and best regards.
Dear Christian,
Thanks for your response and of course the link. I have a question. Can this dye affect the cells adversley? Because I need them alive with full viability for further work.
The brochure says it can be a mutagen...
Regards,
Shahin
Hi, the brochure says it can be a mutagen..., but you will resuse the parasite sample again or will use it just for counting? you can use an hemocytometer for couting.
I think that the cells will remain viable. However, the mutagenicity might pose a problem if you would like to conduct long-term experiments. There might be some changes to the genotype of certain cells (although I think that the dye is only mildly mutagenic). In summary, I think that you won't run into problems because you are probably not working at the single cell level?
The parasites are going to be used in vaccination trials. Therefore, I want to measure the number accurately. If the dye affects the cells at the nucleotide level and change the genotype, then the results will be biased....
... But you will reuse the cells to produce your vaccine ? Or you will collect a sample from a batch for couting the cells and use the rest of the batch for vaccine?
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