RNA analysis is essential for understanding biological activities of a cell or tissue. Unfortunately, retrieval of RNA from existing archives of human temporal bones has proven extremely difficult due to degradation of RNA molecules. The major factors that contribute to degradation of RNA in specimens from autopsied temporal bones are tissue autolysis due to time elapsed before autopsy, and technical problems in processing the bones after harvest. We therefore focused on improving the survival of RNA in human temporal bones by shortening the time to autopsy and through modification of the processing technique by removing targeted tissues directly from the temporal bones and by avoiding time-consuming decalcification and celloidin- embedding. Eight temporal bones collected at immediate autopsies were used in this study. Representative mRNAs, ranging from high (MUC5B, physically unstable) to low (β-actin, physically stable) molecular weights, and from abundant (MUC5B) to non-abundant (MUC1) RNA, were studied by in situ hybridization, Northern blot technique, or both. Using this modified protocol in autopsies performed up to 6 h after death, the existence of mRNAs was demonstrated in all bones studied. This improved method demonstrates the feasibility of the use of autopsied temporal bone tissues for RNA analysis.
Bibliographical noteFunding Information:
We thank JoAnn Knox for editorial assistance during preparation of the manuscript. This research was supported in part by the Multiple Districts (5M) International Lion’s Club and the International Hearing Foundation.
- Eustachian tube
- Middle ear
- RNA analysis
- Temporal bone