Evaluation of Non-Formalin Tissue Fixation for Molecular Profiling Studies
Author(s): Gillespie JW, Best CJ, Bichsel V, Cole K, Greenhut S, Hewitt SM, Ahram M, Gathright Y, Merino M, Strausberg R, Epstein J, Hamilton SR, Gannot G, Baibakova G, Calvert V, Flaig M, Chuaqui RF, Herring J, Pfeifer J, Petricoin EF, Linehan WM, Duray P, Bova G, Emmert-Buck MR
Publication: Am J Pathol, 2002, Vol. 160, Page 449
PubMed ID: 11839565 PubMed Review Paper? No
Purpose of Paper
Conclusion of Paper
Studies
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Study Purpose
To compare the quality and quantity of protein extracted from: 1) tissue fixed in 70% ethanol and embedded in paraffin; 2) tissue fixed in 70% ethanol and embedded in polyester wax; 3) formalin-fixed paraffin-embedded tissue; and 4) snap frozen tissue.
Summary of Findings:
While 1D gel analysis for protein quality and quantity indicated that ethanol-fixed paraffin embedded tissue yielded similar and superior results when compared to snap frozen and formalin-fixed tissue, respectively, observations with 2D-PAGE analysis included a 50% reduction in total protein quantity in ethanol-fixed paraffin embedded tissue when compared to snap frozen controls. As observed in immunoblots, bands corresponding to prostate specific antigen (PSA) were present in ethanol-fixed paraffin embedded tissues and were similar in intensity to snap frozen controls. Immunohistochemistry results for PSA were comparable in paraffin embedded tissues fixed in 70% ethanol and formalin.
Biospecimens
Preservative Types
- Ethanol
- Formalin
- Frozen
Diagnoses:
- Neoplastic - Carcinoma
Platform:
Analyte Technology Platform Protein Immunohistochemistry Protein Western blot Protein 1D/2D gels Pre-analytical Factors:
Classification Pre-analytical Factor Value(s) Biospecimen Preservation Type of fixation/preservation Ethanol
Formalin (buffered)
Snap frozen
Immunohistochemistry Specific Targeted peptide/protein PSA
Western blot Specific Targeted peptide/protein PSA
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Study Purpose
To compare the quality and quantity of RNA extracted from: 1) tissue fixed in 70% ethanol and embedded in paraffin; 2) tissue fixed in 70% ethanol and embedded in polyester wax; 3) formalin-fixed paraffin-embedded tissue; and 4) snap frozen tissue.
Summary of Findings:
Although quality was reduced compared to snap frozen tissue, sufficient RNA was recovered from ethanol-fixed tissue that was paraffin or polyester wax embedded for RT-PCR and cDNA library production. During RNA extraction, incubation at 60 degrees C improved RNA yield of frozen and ethanol-fixed tissues, while incubation at 80 degrees C or greater resulted in fragmentation.
Biospecimens
Preservative Types
- Ethanol
- Frozen
Diagnoses:
- Neoplastic - Carcinoma
Platform:
Analyte Technology Platform RNA RT-PCR RNA Electrophoresis Pre-analytical Factors:
Classification Pre-analytical Factor Value(s) Biospecimen Preservation Embedding medium Paraffin
Polyester wax
RT-PCR Specific RNA incubation temperature 4 degrees C
60 degrees C
80 degrees C
Biospecimen Preservation Type of fixation/preservation Snap frozen
Ethanol
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Study Purpose
To compare the quality and quantity of DNA extracted from: 1) tissue fixed in 70% ethanol and embedded in paraffin; 2) tissue fixed in 70% ethanol and embedded in polyester wax; 3) formalin-fixed paraffin-embedded tissue; and 4) snap frozen tissue.
Summary of Findings:
DNA recovered from ethanol-fixed tissue in comparison to formalin fixation was superior in quality and PCR performance for the microsatellite marker D17S926. PCR performance was similar among ethanol-fixed specimens embedded in either paraffin or polyester wax.
Biospecimens
Preservative Types
- Ethanol
- Frozen
- Formalin
Diagnoses:
- Neoplastic - Carcinoma
Platform:
Analyte Technology Platform DNA Electrophoresis DNA PCR Pre-analytical Factors:
Classification Pre-analytical Factor Value(s) Biospecimen Preservation Type of fixation/preservation Ethanol
Formalin (buffered)
Snap frozen
Biospecimen Preservation Embedding medium Paraffin
Polyester wax
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Study Purpose
The histology of prostate and kidney tissues fixed in: 1) 70% ethanol and embedded in paraffin; 2) 70% ethanol and embedded in polyester wax; 3) formalin and paraffin-embedded; and 4) snap frozen were assessed for quality based on nuclear morphology, cellular morphology, tissue architecture, and staining characteristics.
Summary of Findings:
Ethanol fixation yielded histological quality comparable to formalin fixation and superior to snap freezing for tissue architecture and staining quality. Preservation of nuclear detail was also superior following ethanol compared to formalin fixation. Technical difficulties associated with polyester wax embedding impeded the use of tissue sections larger than 1 cm.
Biospecimens
Preservative Types
- Ethanol
- Formalin
- Frozen
Diagnoses:
- Neoplastic
Platform:
Analyte Technology Platform Morphology H-and-E microscopy Pre-analytical Factors:
Classification Pre-analytical Factor Value(s) Biospecimen Preservation Embedding medium Paraffin
Polyester wax
Biospecimen Preservation Type of fixation/preservation Ethanol
Formalin (buffered)
Snap frozen