[A study on the method of DNA extraction from unbuffered formalin-fixed and paraffin-embedded samples].
OBJECTIVE
Unbuffered formalin is widely used to fix resected specimens in China. The DNA in unbuffered formalin-fixed and paraffin-embedded tissues is usually degraded seriously, so the extraction of DNA from these samples is difficult. This study was conducted to seek an optimal method to extract DNA from these samples.


METHODS
Fifteen blocks of esophageal carcinoma resected in Fourth Hospital of Hebei Medical University in 2000 were selected. The cells were lyzed by proteinase K digestion or heating under different pH values, then DNA was extracted by phenol:chloroform. After that, four parameters (deparaffined by xylene or histolene; digested for 48 h or 72 h at 37 degrees C or 56 degrees C; extracted by salting-out or phenol:chloroform) were optimized according to the principle of cross design. At last, the quality of obtained DNA was analyzed with electrophoresis and PCR amplification.


RESULTS
The quality and quantity of DNA obtained by proteinase K digestion (the average yield is 17.88 microg) were better than that of heating under different pH (7-12)(P< 0.05). The quality and quantity of DNA digested at 56 degrees C were better than that at 37 degrees C, and similarly, digestion for 72 hours was better than that for 48 hours. The methods of deparaffin and extraction had no obvious influence on the quality and quantity of DNA.


CONCLUSIONS
By means of NaCl salting-out after proteinase K digestion, more reliable quality of DNA can be obtained from unbuffered formalin-fixed and paraffin-embedded samples. Furthermore,digestion for three days at 56 degrees C is more likely to obtain DNA with high quality and quantity.
Tian
Zi-Qiang
ZQ
Department of Thoracic Surgery, Fourth Hospital of Hebei Medical University, Shijiazhuang, Hebei, 050011, PR China.
Liu
Jun-Feng
JF
Zhang
Shao-Wei
SW
Li
Bao-Qing
BQ
Wang
Fu-Shun
FS
Zhang
Yue-Feng
YF
D004247:DNA / Q000302:isolation & purification*; D005557:Formaldehyde; D006801:Humans; D006863:Hydrogen-Ion Concentration; D016612:Paraffin Embedding; D016133:Polymerase Chain Reaction; D016707:Tissue Fixation
D005557:Formaldehyde; D004247:DNA