I am using fluorescein to test my sample preparation procedures for qPCR experiments. How should I clean up any possible dye contamination on qPCR or the instrument for sample preparation? Can bleach or ethanol destroy the fluroescein molecule to make it not fluorescent any more?
bleach may, ethanol will not destroy the fluroescein molecule
Ethanol may wash away the fluroescein molecule, water or basic diluent NaOH may wash out the fluroescein molecule better.
May be use bleach, then do water wash well!
Dear Nick,
please tell us (the answerers) if you use the free acid of fluorescein or the sodium salt.Aldrich/Merck sells the 2 dyes.
For a good answer and help, it is important to know the pH of your solution for qPCR. Why? Fluorescein molecule has a pKa/pKb value ratio (see google scholar) and has different physico-chemical properties (e.g. solubility , UV-Vis spectroscopy, fluorescence a.s.o....).
Concerning this part of your question:
Can bleach or ethanol destroy the fluroescein molecule to make it not fluorescent any more?
please read:
https://www.sciencedirect.com/science/article/pii/B9780128156476000108 and
Article Assembly properties of fluorescein-labeled tubulin in vitro ...
please read the cited references in these 2 papers.
Thank you in advance and good luck.
JRG
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