The effect of formalin fixation on DNA and the extraction of high-molecular-weight DNA from fixed and embedded tissues.
The effect of formalin fixation on DNA and extraction of DNA from fixed tissues was investigated to retrieve archival tissue samples stored in pathology departments for molecular biological studies. Aldehyde fixatives resulted in degradation of DNA at room temperature but not at 4 degrees C. The degradation also occurred in formalin when the pH or the salt concentration was low, or the formic acid level was high. Restriction endonuclease digestion of fixed DNA was incomplete after formalin fixation and this was also temperature-dependent. Thus, relatively intact DNA was obtained from the tissues fixed in buffered formalin at 4 degrees C or fixed with microwave irradiation. The use of modified tissue-lysing buffer containing 4M urea allowed extraction of high-molecular-weight DNA suitable for Southern blot analysis from fixed and embedded tissues. In conclusion, fixation with buffered formalin at 4 degrees C permitted extraction of DNA of sufficient quality for Southern blot analysis.
Department of Geriatric Medicine, Faculty of Medicine, Kyoto University, Japan.
M
M
Koshiba
K
K
Ogawa
S
S
Hamazaki
T
T
Sugiyama
O
O
Ogawa
T
T
Kitajima
D000447:Aldehydes / Q000494:pharmacology; D000818:Animals; D010582:Bacteriophage lambda / Q000235:genetics; D004247:DNA / Q000187:drug effects* / Q000302:isolation & purification / Q000528:radiation effects / Q000648:ultrastructure; D004249:DNA Damage; D004262:DNA Restriction Enzymes / Q000494:pharmacology; D005557:Formaldehyde / Q000494:pharmacology*; D008099:Liver; D051379:Mice; D008872:Microwaves; D012965:Sodium Chloride / Q000494:pharmacology*; D016707:Tissue Fixation / Q000379:methods*
D000447:Aldehydes; D005557:Formaldehyde; D012965:Sodium Chloride; D004247:DNA; D004262:DNA Restriction Enzymes