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JC virus genotyping using formalin-fixed, paraffin-embedded renal tissues
Hiroshi Ikegaya1, Hirotaro Iwase, Huai-Ying Zheng
1Department of Forensic Medicine, Graduate School of Medicine, University of Tokyo, 7-3-1 Hongo, Tokyo 113-0033, Japan. ikegaya-tky@umin.ac.jp
Journal of Virological Methods
|April 26, 2005
Summary
Formalin fixation can alter JC virus (JCV) DNA sequences in kidney tissue, potentially affecting geographic origin identification. Frozen or briefly formalin-fixed samples are recommended for accurate JCV DNA genotyping.
Area of Science:
- Forensic Science
- Molecular Biology
- Virology
Background:
- JC virus (JCV) DNA genotyping in renal tissue aids in identifying the geographic origin of unidentified cadavers.
- Previous studies utilized fresh or frozen autopsy samples for JCV DNA analysis.
- The impact of formalin fixation on JCV DNA sequencing in kidney tissue requires investigation for broader sample applicability.
Purpose of the Study:
- To evaluate the effects of formalin fixation on JC virus (JCV) DNA sequences in kidney tissue.
- To determine the suitability of formalin-fixed, paraffin-embedded (FFPE) samples for JCV DNA genotyping.
- To assess the reliability of JCV DNA genotyping from FFPE samples for forensic applications.
Main Methods:
- JC virus (JCV) DNA was amplified from renal tissues stored for one year under three conditions: frozen (-80°C), formalin-fixed paraffin-embedded (FFPE), and 5% formalin-soaked.
- A 610 bp typing region of the JCV genome was PCR-amplified from all sample types.
- Amplified fragments were cloned and sequenced to analyze sequence modifications.
Main Results:
- Frozen samples yielded single, 'original' JCV DNA sequences.
- FFPE samples revealed original sequences alongside variants with 1-6 base substitutions.
- Formalin-soaked samples showed original sequences plus variants with 1-5 and 10-13 base substitutions; significant substitutions (10-13) caused a shift in observed JCV genotypes.
Conclusions:
- Minor base substitutions (1-6) in formalin-fixed samples do not impede JCV DNA genotyping.
- Extensive base substitutions (10-13) resulting from prolonged formalin exposure can alter JCV genotype determination.
- Accurate JCV DNA genotyping for forensic purposes is feasible with frozen specimens or samples with short-term formalin fixation.