Abstract

BUSTAMANTE, JAVIER ANDRÉS; ASTUDILLO, MIRYAM; PAZOS, ALVARO JAIRO  and  BRAVO, LUIS EDUARDO. Evaluation of Two Methods DNA Extraction from Formalin-Fixed, Paraffin-Embedded Tissues on Non-Optimal Conditions. Acta biol.Colomb. [online]. 2011, vol.16, n.2, pp.83-98. ISSN 0120-548X.

Paraffin wax embedded tissues are an invaluable material for retrospective studies requiring the application of molecular analysis. Multiple methods are available to extract DNA from these kind of samples. However, the most common methods are slow and the reagents often contribute to the fragmentation of genetic material. In order to optimize the procedure, two methods for DNA extraction from paraffin embedded tissue non-optimal conditions were used. 47 blocks containing paraffin-embedded biopsies of pleura, lung and pericardium from 24 patients (66.6% males) older than 18 years, with biopsy proven chronic granulomatous inflammation referred to the Department of Pathology at University hospital of Valle between 2002 and 2007 were selected. Each sample was subjected to 10 cuts and was to two methods of DNA extraction: 1. conventional and 2. QIAamp-DNA mini kit®. The efficiency of the extracted DNA, was assessed by spectrophotometry and PCR amplification of a fragment of the housekeeping gene GAPDH. The concentration of DNA samples extracted by the conventional method was of 65.52 ng/µL ± 11.47 (mean ± SE) and the 260/280 absorbance ratio ranged between 0.52 and 2.30 the average concentration of DNA of the samples extracted by the commercial method was 60.89 ng/µL ± 6.02 (mean ± SE), with an absorbance that fluctuated between 0 and 2.64. The DNA obtained was amplified by PCR, of 47 samples extracted by both methods, 25 and 23 respectively the GAPDH gene amplified successfully. The methods used to obtain DNA showed similar performance, highlighting the potential utility of both extraction methods for the retrospective studies from paraffin embedded tissues in unsuitable conditions.

Keywords : biopsy; granuloma; nucleic acids; DNA fragmentation; polymerase chain reaction..

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