Extraction of DNA from paraffin blocks for Southern blot analysis.
Tissues stored as paraffin blocks are a potential source of DNA for retrospective clinicogenetic analysis. To assess the feasibility of Southern blot analysis, DNA extracted from paraffin blocks was compared with DNA obtained from fresh-frozen controls of the same tissues. Sections 50-100 microns thick cut from paraffin blocks of 11 normal tissues, 18 lymphoid lesions, and 9 gastric carcinoma samples were deparaffinized and incubated at 45 degrees C for 48 to 72 h in a sodium dodecyl sulfate (SDS)/proteinase K solution. Following organic extraction, alcohol precipitation, restriction endonuclease digestion, and gel electrophoresis, DNA was transferred to nylon membranes. 32P-labelled DNA probes for the immunoglobulin heavy-chain locus and T-cell receptor beta-chain gene were hybridized to the normal tissue and lymphoid samples; the gastric cancers were probed for the HER-2/neu protooncogene. Intact DNA was obtained from the majority of formalin-fixed samples, yielding results qualitatively similar to those from fresh tissues. Degradation is the most significant problem in analyzing DNA extracted from paraffin blocks and compromises accurate quantitation. DNA analysis using paraffin-embedded tissue has potential clinical and research applications and may be a particularly useful way to study gene abnormalities in unusual tumors infrequently available as fresh specimens.
Mies
C
C
Department of Pathology, Memorial Hospital, New York, New York 10021.
Houldsworth
J
J
Chaganti
R S
RS
D001345:Autoradiography; D015139:Blotting, Southern; D004247:DNA / Q000032:analysis*; D005615:Freezing; D006801:Humans; D009693:Nucleic Acid Hybridization; D010232:Paraffin; D011309:Preservation, Biological / Q000379:methods*
D010232:Paraffin; D004247:DNA