The analysis of nucleic acid from formalin-fixed, paraffin-embedded (FFPE) specimens is challenging due to the extensive cross-linking of all tissue components during the fixation process. These challenges include chemical modification of the DNA, cross-linking of DNA with other molecules, degradation of the DNA, and the limited amount of nucleic acid in the samples.
The QuickExtract™ FFPE DNA Extraction Kit is a fast, simple, and inexpensive method for preparing genomic DNA for PCR amplification from archival samples (Fig. 1). The protocol requires only heat treatment to melt the paraffin, lyse the cells, decrease the fomalin-induced cross-linking in the sample, and degrade compounds inhibitory to amplification. Following heat treatment, the sample DNA is ready for PCR (Figs. 2 and 3).
Note: This product is only intended for PCR- and qPCR-based applications. If DNA is required for other molecular biology applications, consider the MasterPure™ DNA Purification Kit.
Figure 1. Overview of the QuickExtract™ FFPE DNA extraction procedure.
Figure 2. PCR amplification of DNA from a slide-mounted, FFPE preserved human skeletal muscle tissue section, extracted with the QuickExtract™ FFPE DNA Extraction Kit. Two microliters of undiluted, extracted DNA was amplified with primers for three different loci: tumor protein 53 (TP53), dystrophin (DMD), and tumor necrosis factor (TNF). The products were separated on a 3% agarose gel and were visualized with SYBR® Gold. Lane M, 100-bp DNA ladder; lane 1, exon 2 of TP53; lane 2, exon 3 of TP53; lane 3, exon 11 of TP53; lane 4, exon 6 of DMD; lane 5, exon 50 of DMD; lane 6, exon 3 of DMD; lane 7, exon 4 of TNF.
Figure 3. PCR amplification of DNA from slide-mounted, FFPE-preserved, human tissue sections, combined and extracted with the QuickExtract™ FFPE DNA Extraction Kit. Two microliters of undiluted, extracted DNA was amplified with primers for seven different STR loci. The products were separated on a 3% agarose gel and were visualized with SYBR® Gold. The expected allele-to-allele variation in STR sequences results in multiple products per primer set. Lane M, 100-bp DNA ladder; lane 1, RENA4; lane 2, D3S1358; lane 3, D7S820; lane 4, THO1; lane 5, D19S253; lane 6, D21S11; lane 7, AMEL.