Hi All,
I am looking for a reagent/buffer which could allow me to stabilize intestinal tissue just after dissecting mice.
We will be dissecting several mice in a day, and if we keep flushed cleaned intestines in DMEM+5% FBS on ice (5-6 hours) we get low viability of epithelial cells (30-40%) - that we get after doing EDTA and enzymatic digestion; Dispase + DNAase on intestines. And, because of poor viability i get poor quality RNA.
Briefly, I intend to prepare single epithelial-cell suspension from the stabilized tissue, do sorting and extract RNA from the sorted cells.
So, ideally the stabilizing buffer i am looking for should
- be able to stabilize RNA with in the tissue
- not interfere with the EDTA + Enzyme digestion (of course, i will wash intestines thoroughly before i proceed to the digestion steps)
If anybody knows about such a reagent , please please let me know.
Thanks in advance.
Commercially available platforms to stabilize messenger RNA (mRNA) and microRNA are critically designed to optimize and ensure the quality and integrity of . One such reagent is RNAsecure resuspension solution which when introduced to a buffer or even purified RNA pellets can inactivate RNAses.
Obtaining high-quality, intact RNA is the first, and often most critical, step in performing gene expression analysis. In order to isolate high-quality RNA, the tissue has to be either: a) processed immediately after harvest, b) snap-frozen, or c) stabilized in an intermediary solution to preserve RNA integrity and allow for storage.
Regards