title: PCR amplification and high throughput sequencing of immunoglobulin heavy chain genes from formalin-fixed paraffin-embedded human biopsies.
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.
authors.affiliation: The Mina and Everard Goodman Faculty of Life Sciences, Bar-Ilan University, Ramat Gan, Israel. hillatk73@gmail.com
authors.forename: Hilla
authors.initials: H
authors.lastname: Tabibian-Keissar
authors.forename: Ginette
authors.initials: G
authors.lastname: Schibby
authors.forename: Miri
authors.initials: M
authors.lastname: Michaeli
authors.forename: Aviya
authors.initials: A
authors.lastname: Rakovsky-Shapira
authors.forename: Noemie
authors.initials: N
authors.lastname: Azogui-Rosenthal
authors.forename: Deborah K
authors.initials: DK
authors.lastname: Dunn-Walters
authors.forename: Kinneret
authors.initials: K
authors.lastname: Rosenblatt
authors.forename: Ramit
authors.initials: R
authors.lastname: Mehr
authors.forename: Iris
authors.initials: I
authors.lastname: Barshack
mesh_terms: D000328:Adult; D000368:Aged; D005260:Female; D005557:Formaldehyde; D050438:Genes, Immunoglobulin Heavy Chain; D014644:Genetic Variation; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D007143:Immunoglobulin Heavy Chains / Q000235:genetics*; D008297:Male; D008875:Middle Aged; D016612:Paraffin Embedding; D016133:Polymerase Chain Reaction; D011947:Receptors, Antigen, B-Cell / Q000235:genetics; D017422:Sequence Analysis, DNA; D016707:Tissue Fixation
chemical_list: D007143:Immunoglobulin Heavy Chains; D011947:Receptors, Antigen, B-Cell; D005557:Formaldehyde