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Man, YG; Moinfar, F; Bratthauer, GL; Kuhls, EA; Tavassoli, FA.
An improved method for DNA extraction from paraffin sections.
Pathol Res Pract. 2001; 197(9):635-642 Doi: 10.1078/0344-0338-00138
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Co-authors Med Uni Graz: Moinfar Farid
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Abstract:: The acquisition of comparable quality and quantity of DNA extracts is the prerequisite to the success of comparative genetic analyses. Although several DNA extracting protocols on paraffin sections have been introduced, the importance of deparaffinization, the procedure for obtaining an adequate hematoxylin staining, the significance of the ratio of the cell number to the enzyme volume, and a practical means for monitoring the digestion process have not been sufficiently addressed. These, however, are the most important factors accountable for a failure of DNA extraction. To minimize the impact of these factors, we have developed several unique strategies, including: (1) incubating sections at 80 degrees C for 30-60 minutes prior to xylene treatment, (2) checking each section to insure the complete removal of paraffin; (3) treating hematoxylin stained sections or cells with de-staining solutions; (4) using a micrometer inserted into the eyepiece of a microscope to estimate the number of cells collected and adjusting the enzyme volume according to the cell number; and (5) monitoring the digestion process with a magnifier. With these strategies, we have been able to consistently obtain comparable quality and quantity of DNA extracts which yielded uniform PCR products regardless of variations in tissue embedding and processing.
Find related publications in this database (using NLM MeSH Indexing): Clone Cells -; DNA - isolation and purification; Dissection - isolation and purification; Female - isolation and purification; Humans - isolation and purification; Loss of Heterozygosity - isolation and purification; Micromanipulation - isolation and purification; Nucleic Acid Hybridization - isolation and purification; Paraffin - chemistry; Paraffin Embedding - chemistry; Staining and Labeling - chemistry
Find related publications in this database (Keywords): histotechniques; paraffin sections; DNA extraction; comparative genetic analysis