Method for the purification of tissue DNA suitable for PCR after fixation with Bouin's fluid. Uses and limitations in microsatellite typing.
Paraffin-embedded tissues are often the only available material to perform polymerase chain reaction (PCR)-based analysis in various medical purposes. Unfortunately, the use in many countries of acid fixatives such as Bouin's fluid limits the use of such a material for molecular analysis. This article reports the methodological details of a DNA purification technique from Bouin-fixed and paraffin-embedded samples based on a double washing, in an alcohol then in an aqueous medium, of the DNA, which enables PCR reactions from this material. Comparison of the results with those obtained by organic solvent purification of DNA from frozen tissue fragments showed excellent reproducibility in terms of detection of an amplification product on agarose gel. However, differences between the methods were quite frequently seen in the allelic typing profile of microsatellite sequences (CA repeats), either as neo-alleles or by the loss of normal alleles in the fixed materials that constitute a limitation in using DNA from Bouin-fixed tissue as a substrate for fine allelotyping.
Longy
M
M
Laboratory of Molecular Oncology, Institut Bergonie, Bordeaux, France.
Duboue
B
B
Soubeyran
P
P
Moynet
D
D
D019342:Acetic Acid; D000483:Alleles; D002869:Chromosome Aberrations; D015925:Cryopreservation; D004273:DNA, Neoplasm / Q000302:isolation & purification*; D004587:Electrophoresis, Agar Gel; D005404:Fixatives; D005557:Formaldehyde; D006801:Humans; D018895:Microsatellite Repeats; D009369:Neoplasms / Q000235:genetics / Q000473:pathology; D016612:Paraffin Embedding; D010851:Picrates; D016133:Polymerase Chain Reaction; D016707:Tissue Fixation / Q000379:methods
D004273:DNA, Neoplasm; D005404:Fixatives; D010851:Picrates; D005557:Formaldehyde; C026239:Bouin's solution; D019342:Acetic Acid