PCR amplification and high throughput sequencing of immunoglobulin heavy chain genes from formalin-fixed paraffin-embedded human biopsies

Hilla Tabibian-Keissar, Ginette Schibby, Miri Michaeli, Aviya Rakovsky-Shapira, Noemie Azogui-Rosenthal, Deborah K. Dunn-Walters, Kinneret Rosenblatt, Ramit Mehr, Iris Barshack

Research output: Contribution to journalArticlepeer-review

Abstract

The use of high throughput sequencing (HTS) technologies in biomedicine is expanding in a variety of fields in recent years. The 454 system is an HTS platform that is ideally suited to characterize B cell receptor (BCR) repertoires by sequencing of immunoglobulin (Ig) genes, as it is able to sequence stretches of several hundred nucleotides. Most studies that used this platform for antibody repertoire analyses have started from fresh or frozen tissues or peripheral blood samples, and rely on starting with optimal quality DNA. In this paper we demonstrate that BCR repertoire analysis can be done using DNA from formalin-fixed paraffin-embedded (FFPE) human tissue samples. The heterogeneity of BCR repertoires we obtained confirms the plausibility of HTS of DNA from FFPE specimens. The establishment of experimental protocols and computational tools that enable sequence data analysis from the low quality DNA of FFPE tissues is important for enabling research, as it would enable the use of the rich source of preserved samples in clinical biobanks and biopsy archives.

Original languageEnglish
Pages (from-to)182-187
Number of pages6
JournalExperimental and Molecular Pathology
Volume94
Issue number1
DOIs
StatePublished - Feb 2013

Keywords

  • FFPE tissues
  • Formalin-fixed paraffin-embedded tissues
  • High throughput sequencing
  • HTS
  • Ig
  • Ig variable region genes
  • MID
  • PCR
  • SHM

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