Biobanking of fresh frozen tissue: RNA is stable in nonfixed surgical specimens
Author(s): Micke P, Ohshima M, Tahmasebpoor S, Ren ZP, Ostman A, Pontén F, Botling J
Publication: Lab Invest, 2006, Vol. 86, Page 202
PubMed ID: 16402036 PubMed Review Paper? No
Purpose of Paper
Conclusion of Paper
Studies
-
Study Purpose
To determine if storage media, temperature, and the length of time prior to freezing in OCT impact RNA integrity using the following experimental paradigm: fresh tonsil and colon tissue were 1) left at room temperature, 2) stored on ice, 3) stored in cold normal saline, or 4) stored in cold RNAlater prior to being snap frozen after 0.5, 1, 3, 6 and 16 h durations.
Summary of Findings:
Microchip electrophoresis of RNA yielded distinct 28S and 18S ribosomal peaks for all samples and storage conditions. Ribosomal 28S/ 18S ratios decreased as storage time increased for all conditions, although storage in saline for 16 h alone differed significantly from 0 h controls.
Biospecimens
Preservative Types
- OCT
- RNAlater
- Other Preservative
Diagnoses:
- Normal
Platform:
Analyte Technology Platform RNA Electrophoresis Pre-analytical Factors:
Classification Pre-analytical Factor Value(s) Storage Storage temperature 0 degrees C
4 degrees C
22 degrees C
Biospecimen Preservation Type of fixation/preservation Normal saline
None
RNAlater
Biospecimen Acquisition Cold ischemia time 0 h
0.5 h
1 h
3 h
6 h
16 h
-
Study Purpose
To assess RNA integrity in numerous normal and malignant tissue samples stored on wet ice (15 min- 3 h) prior to processing for inclusion in a biobank.
Summary of Findings:
Clear 28S and 16S ribosomal peaks were observed in 96% of the 47 samples examined, indicating intact RNA. RNA integrity did not appear to be dependent on diagnosis or time to freezing.
Biospecimens
Preservative Types
- OCT
Diagnoses:
- Neoplastic - Other
- Normal - Normal Adjacent
Platform:
Analyte Technology Platform RNA Electrophoresis Pre-analytical Factors:
Classification Pre-analytical Factor Value(s) Biospecimen Acquisition Cold ischemia time 15 min
3 h
-
Study Purpose
To determine if storage media and length of time prior to freezing impact the preservation of cellular structure as observed in hematoxylin and eosin stained tissue sections.
Summary of Findings:
Preservation of cellular structure was only affected in samples stored in RNAlater, which displayed evidence of tissue condensation, cell shrinkage, and hyperchromatic nuclei.
Biospecimens
Preservative Types
- OCT
- Other Preservative
- RNAlater
Diagnoses:
- Normal
Platform:
Analyte Technology Platform Morphology H-and-E microscopy Pre-analytical Factors:
Classification Pre-analytical Factor Value(s) Biospecimen Preservation Type of fixation/preservation RNAlater
OCT
Normal saline
Biospecimen Acquisition Cold ischemia time 0 h
0.5 h
1 h
3 h
6 h
16 h
Storage Storage temperature 0 degrees C
4 degrees C
22 degrees C
-
Study Purpose
To determine if storage media and the length of time prior to freezing impact the expression of representative immediate early (cfos), growth factors (TGFbeta1), signal transduction (SMAD7), hypoxia responsive (HIF1alpha), apoptosis regulatory (Bcl2), and cell cycle regulatory (PCNA) genes as measured by real time quantitative RT-PCR.
Summary of Findings:
Expression of targeted genes in samples stored on ice for 16 h (but not earlier timepoints) differed significantly from 0 h controls, as observed by decreased HIF1alph, TGFbeta1, and Bcl2 levels. Samples stored at room temperature for 6 h or more exhibited a robust decrease in cfos, while all other examined genes remained stable. Storage in normal saline after only 30 min yielded increased cfos and SMAD7 expression, as well as HIF1alpha, Bcl-2, and PCNA after 16 h. Samples stored in RNAlater exhibited the greatest and most variable differences in gene expression: after storage from 0.5 to 16 h levels of cfos, TGFbeta1, Bcl2, and SMAD7 markedly increased.
Biospecimens
Preservative Types
- OCT
- RNAlater
- Other Preservative
- None (Fresh)
Diagnoses:
- Normal
Platform:
Analyte Technology Platform RNA Real-time qRT-PCR Pre-analytical Factors:
Classification Pre-analytical Factor Value(s) Storage Storage temperature 4 degrees C
0 degrees C
22 degrees C
Storage Storage duration 0.5 h
1h
3 h
6 h
16 h
Biospecimen Preservation Type of fixation/preservation Normal saline
None
RNAlater
OCT
Real-time qRT-PCR Specific Targeted nucleic acid c-fos
TGF-beta 1
SMAD7
HIF-1 alpha
Bcl-2
PCNA