Hello all,
We have been trying to isolate RNA from whole blood using different kit methods (Promega and others) but we get degraded RNA every time. Please tell us any manual method like Trizol and CTAB method in which we do not need to extract buffy coat first. Also indicate the homogenization procedure. Is it true that haemoglobin binds RNA and precipitates with it so that is why we are not getting RNA?
Many thanks
I have used the Paxgene System (tube and kit) to isolate RNA from whole blood (2.5 mL per sample). Both integrity (RIN>8.0) and yield (up to 10 ug of total RNA) have presented good results. However, I've noticed that samples from individuals submitted to some stress (e.g. disease, drug treatment) presented a lower integrity and yield compared to healthy samples, but still reasonable results have been obtained.
RiboPure™ RNA Purification Kit for blood (Life Technologies) works very well. Trizol LS Reagent (also Life Technologies) also works well. Here it is the protocol: https://tools.lifetechnologies.com/content/sfs/manuals/trizol_ls_reagent.pdf
U can use TRI reagent which makes the process very easy for isolating RNA. But before using TRI reagent, I would suggest to purify WBC (buffy coat) by ficoll 400.
Using a kit has a big disadvantage of filters with lower (binding capcacity)capacity to hold nucleic acid and so u might not get concentrations more than 100 ng/µl. Anything more than the capacity of the filter, will be washed out with waste.
I suggest you to use a Life Technology kit. But because each RNA extraction kit from blood samples is optimized for something, i suggest you to take a look at this page and choose the best one for your aim.
http://www.lifetechnologies.com/it/en/home/life-science/dna-rna-purification-analysis/rna-extraction/rna-sample-extraction/rna-isolation-from-blood.html
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