Simplified procedures for applying the polymerase chain reaction to routinely fixed paraffin wax sections.
Author(s): Coates PJ, d'Ardenne AJ, Khan G, Kangro HO, Slavin G
Publication: J Clin Pathol, 1991, Vol. 44, Page 115-8
PubMed ID: 1650795 PubMed Review Paper? No
Purpose of Paper
Conclusion of Paper
Studies
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Study Purpose
The purpose of this study was to compare PCR efficiency for different size amplicons when varying the number and thickness of paraffin sections used for DNA extraction, and inclusion of a deparaffinization step.
Summary of Findings:
PCR amplification (at 50 cycles) of the 110 bp beta-globin fragment was successful in 100% of FFPE specimens, while the 355 bp fragment was amplified in 88% of specimens. PCR performance was not affected by the size or quantity of paraffin sections or whether a separate deparaffinization step was performed. PCR amplification of a 153 bp Epstein Barr virus fragment was successful in all 9 FFPE specimens examined.
Biospecimens
Preservative Types
- Formalin
Diagnoses:
- Neoplastic - Carcinoma
Platform:
Analyte Technology Platform DNA PCR Pre-analytical Factors:
Classification Pre-analytical Factor Value(s) Analyte Extraction and Purification Deparaffinization Xylene
None
Biospecimen Aliquots and Components Aliquot size/volume One 5 um section
Four 10 um sections
PCR Specific Length of gene fragment 110 bp
153 bp
355 bp
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Study Purpose
The purpose of this study was to assess whether a two-step PCR protocol would reduce formation of primer-dimer PCR artifacts.
Summary of Findings:
The authors note that a 2 step PCR approach, of 25 cycles each, reduced primer-dimer artifacts without adversely impacting amplicon abundance.
Biospecimens
Preservative Types
- Formalin
Diagnoses:
- Neoplastic - Carcinoma
Platform:
Analyte Technology Platform DNA PCR Pre-analytical Factors:
Classification Pre-analytical Factor Value(s) PCR Specific Technology platform One-stage PCR
Two-stage PCR