The Impact of Different Preservation Conditions and Freezing-Thawing Cycles on Quality of RNA, DNA, and Proteins in Cancer Tissue.
BACKGROUND
High-quality cancer tissues are essential for future research, especially molecular research. For the sake of better quality of tissues, some storage methods are chosen according to lab conditions. But the impact of different storing conditions on the quality of RNA, DNA (especially the degree of DNA methylation), and protein of tissues that have undergone a thawing process, is not clear.


METHODS
We analyzed the influence of different storage conditions including in RNALater solution, normal saline, Opti-mum Cutting Temperature compound (OCT), and snap frozen with no protective reagent (as control) in paired tissue samples on the quality of RNA (RNA Integrity Number value and mRNA expression), DNA quality (DNA amplification and DNA methylation degree of gene RASSF1a), and protein quality. Further, we analyzed the RNA quality of tissues that underwent three freeze-thaw cycles.


RESULTS
The RNALater-treated group retained good RNA quality as expected on three repeated freeze-thaw cycles (RIN>8), but the snap-frozen tissues showed relatively poor results after one freeze-thaw cycle (RIN<7) and three times repeated freeze-thaw cycles (RIN<6). RNA from saline- and OCT-treated groups also yielded good results when we repeated freezing and thawing one time (RIN>7) and two times (RIN>6). The impact of different storing conditions on DNA amplification is small. However, DNA methylation and protein quality are different with different storing conditions. OCT seems to be more secure and stable compared with other two experimental groups, and show a similar trend with control group.


CONCLUSIONS
In consideration of budget and efficiency, we suggest OCT as the best storing method that not only preserves RNA quality during the freezing-thawing process well, but also ensures more secure and stable DNA and protein.
1 Laboratory of Molecular Diagnosis of Cancer, West China Hospital, Sichuan University , Chengdu, China .|2 State Key Laboratory of Biotherapy and Cancer Center, West China Hospital, Sichuan University , and Collaborative Innovation Center for Biotherapy, Chengdu, China .
Yaogeng
Y
Wang
1 Laboratory of Molecular Diagnosis of Cancer, West China Hospital, Sichuan University , Chengdu, China .
Hong
H
Zheng
1 Laboratory of Molecular Diagnosis of Cancer, West China Hospital, Sichuan University , Chengdu, China .
Jie
J
Chen
1 Laboratory of Molecular Diagnosis of Cancer, West China Hospital, Sichuan University , Chengdu, China .
Xiaorong
X
Zhong
1 Laboratory of Molecular Diagnosis of Cancer, West China Hospital, Sichuan University , Chengdu, China .
Yu
Y
Wang
1 Laboratory of Molecular Diagnosis of Cancer, West China Hospital, Sichuan University , Chengdu, China .
Zhu
Z
Wang
1 Laboratory of Molecular Diagnosis of Cancer, West China Hospital, Sichuan University , Chengdu, China .|2 State Key Laboratory of Biotherapy and Cancer Center, West China Hospital, Sichuan University , and Collaborative Innovation Center for Biotherapy, Chengdu, China .
Yanping
Y
Wang
D015925:Cryopreservation / Q000379:methods*; D004247:DNA / Q000737:chemistry*; D006801:Humans; D009369:Neoplasms / Q000737:chemistry*; D011506:Proteins / Q000737:chemistry*; D012313:RNA / Q000737:chemistry*; D013696:Temperature
D011506:Proteins; D012313:RNA; D004247:DNA