Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2003 Jun;56(3):184-6.
doi: 10.1136/mp.56.3.184.

PCR analysis in archival postmortem tissues

S Bonin  1 , F Petrera, B Niccolini, G Stanta
Affiliations

PCR analysis in archival postmortem tissues

S Bonin et al. Mol Pathol. 2003 Jun.

Abstract

Background: Formalin fixed and paraffin wax embedded tissues of necropsy origin are an important source for molecular analysis especially in rare diseases, neuropathology, or molecular epidemiology studies. Because of DNA degradation, only short sequences can be amplified from this type of tissue, very often less than 100 bases. This poses problems because studies on polymorphism and mutations occurring in large genes often require the analysis of long sequences.

Methods: The development of a simple treatment to obtain longer fragments of DNA for the analysis of archival postmortem paraffin wax embedded tissues.

Results: It was possible to amplify longer sequences ranging up to 300 bases from postmortem tissues, with no modification to the usual DNA extraction procedures. To obtain longer stretches of DNA, a pre-PCR restoration treatment was required, by filling single strand breaks, followed by a vigorous denaturation step.

Conclusions: The development of this simple treatment allowed the analysis of longer fragments of DNA obtained from archival postmortem paraffin wax embedded tissues.

PubMed Disclaimer

Figures

Figure 1
Figure 1
PCR products for apolipoprotein E (ApoE) analysis in paraffin wax embedded tissues of necropsy and biopsy origin. (A) Postmortem DNA analysis without the denaturation step. Molecular size marker (lane 1), postmortem DNA cases with the restoration step (lanes 2, 4, 6, and 8), and the same postmortem DNA cases without the restoration step (lanes 3, 5, 7, and 9). (B) Biopsy DNA analysis without the denaturation step. Positive control (lane 1), biopsy DNAs with the restoration step (lanes 2, 4, and 6), the same biopsy DNAs without the restoration step (lanes 3, 5, and 7), and the negative control (lane 8). (C) Postmortem sample analysis with the denaturation step. Molecular size marker (lane 1), postmortem samples with the restoration and denaturation steps (lanes 2, 4, 6, and 8), the same postmortem samples without the restoration step (lanes 3, 5, 7, and 9). (D) Biopsy sample analysis with the denaturation step. Positive control (lane 1), biopsy samples with the restoration and denaturation steps (lanes 2, 4, and 6), the same samples without the restoration step (lanes 3, 5, and 7), and the negative control (lane 8).
Figure 2
Figure 2
PCR products in paraffin wax embedded tissues of necropsy and biopsy origin. (A) Postmortem sample analysis for the TTR gene (291 bases). Size marker (lane 1), postmortem samples with the treatment and denaturation steps (lanes 2–10). (B) Biopsy sample analysis for the TTR gene (291 bases). Biopsy samples with the treatment and denaturation steps (lanes 2, 4, and 6), the same biopsy samples without treatment (lanes 1, 3, and 5), the positive control (lane 7), and the negative control (lane 8). (C) Postmortem sample analysis for the TTR gene (339 bases). Molecular size marker (lane 1), postmortem samples with the treatment and denaturation steps (lanes 2–10). (D) Biopsy sample analysis for TTR gene (339 bases). Biopsy samples with the treatment and denaturation steps (lanes 2, 4, and 6), the same biopsy samples without treatment (lanes 1, 3, and 5), the positive control (lane 7), and the negative control (lane 8).
See this image and copyright information in PMC

References

    1. Gressens P, Langston C, Mitchell WJ, et al. Detection of viral DNA in neonatal herpes encephalitis autopsy tissues by solution-phase PCR: comparison with pathology and immunohistochemistry. Brain Pathol 1993;3:237–50. - PubMed
    1. Stanta G, Croce LS, Bonin S, et al. Cohort effect of HCV infection in liver cirrhosis assessed by a 25 year study. J Clin Virol 2000;17:51–6. - PubMed
    1. Lehmann U, Kreipe H. Real-time PCR analysis of DNA and RNA extracted from formalin-fixed and paraffin-embedded biopsies. Methods 2001;25:409–18. - PubMed
    1. Voet D, Voet JG, Pratt CW. Nucleic acid structure. In: Fundamentals of biochemistry. New York: John Wiley and Sons, 1999:739–42.
    1. Ninet B, Rutschmann O, Burkhardt K, et al. Detection of mycobacterial nucleic acids by polymerase chain reaction in fixed tissue specimens of patients with human immunodeficiency virus infection. Swiss HIV cohort study. Diagn Mol Pathol 1999;8:145–51. - PubMed

Publication types