I am aware that whole blood is not great for extracting RNA. But basically i need dto do it. I have found a basic protocol for the trizol method with whole blood however im not convinced my results are correct when analysing on nanodrope. Any advice?
Dear Danielle
If You follow the established protocol, I would pay attention to blood sample collection. That can be a critical step, because RNA can degrade before extraction.
I also would keep in mind that mature erythrocytes, which is a big part of whole blood, do not contain much nucleic acids (no nucleus). So perhaps some kind of enrichment (of, probably, leucocytes) is needed before extraction to get a good amount of RNA.
Thank you. Basically i have been given a cohort of lots of blood samples pre and post delirium. Im doing an investigation involving qpcr. But the people have frozen the samples ! And Without any protectant! So im trying to make the best of a bad situation. So isolating pmbcs first may not work as freezing lyses the cells yes? So i was just going straight to trizol methol from the thawed blood and hoping for the best. Any advice is welcomed!
Dear Danielle
Uf, in this case it can be tough...
Only one thing comes to my mind - add TRIzol to the frozen samples (and add maybe 5 volumes of TRIzol, if it is possible in a given tube) and let blood thaw being in TRIzol. This can be not so easy as TRIzol can initially get frozen if the tube is from LN2, but slowly it should work, I think. Although I am not sure that it will work in respect to final good RNA extraction. The current nano drop results, in my mind, suggest basically no RNA. BTW, do not forget to switch instrument to RNA measurement, as RNA uses smaller coefficient compared to DNA.
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