I want to store the isolated PBMCs of my study subjects in the best possible way that their RNA is preserved and in pristine state for microarray/NXG sequencing analysis.
I tried snap freezing the pelleted PBMC in liquid nitrogen and storing it at -80 degrees.
I also tried storing them in RNA later from Sigma aldrich at 4 degrees, -20 and - 80 degrees centigrade.
But in none of these samples, I was able to achieve a RIN (RNA Integrity Number) above 5 which is a prerequisite for microarray.
Can someone suggest some ways to store isolated PBMCs in an appropriate way to be used for microarray analysis?
If you have the possibility to do so, I would rather recommend to extract RNA from your samples as fast as possible and then to stock your RNAs. I experienced many troubles when throwing a piece of tissue (brain) in RNA later and performing qPCR after extracting RNAs. If this is not possible, I would rather freeze your PBMC pellets slowly, first at -20°C (starting from a ice-cold sample) and then at -80°C if you perform RNA extraction a long time after getting your samples (more than one week). Many people have the rather false impression that RNAs are very fragile and that they will degrade instantly, but believe me it is not the case (and the same is true about RNAses, they are not as aggressive as many think!).
Hope this will help, Fabrice.
I have tried dissolving the PBMC pellet in Trizol and storing it in -20/-80°C for more than a week. Extracted the RNA using chloroform-isoproponol method. Got RIN values as good as 8-9.
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