Abstract
Little is known about molecular testing on tumor tissue retrieved from stained sections, for which there may be a clinical need. We retrospectively analyzed 112 sections from 56 tumor patients using either fluorescence in situ hybridization (FISH) with different probes (19 sections from 17 patients) or Sanger or targeted next generation sequencing for detection of BRAF, EGFR, KRAS, C-KIT, and TP53 mutations (93 sections from 39 patients). Tumor tissue sections had been stained by hematoxylin and eosin (H&E) (42 sections) or by immunohistochemistry for cytoplasmic or nuclear/nuclear-cytoplasmic markers (70 sections) with a peroxidase (P-IHC, with 3,3′-diaminobenzidine as chromogen) or alkaline phosphatase label (AP-IHC, with Warp Red™ as chromogen). For FISH analysis, the concordance rate between the original diagnosis and that obtained on H&E- or P-IHC-stained tissue sections (AP-IHC was not on record for this set of patients) was 95 % (18 out of 19 tumor sections). Only one tumor sample, diffusely positive for MLH1, did not yield any nuclear hybridization signal. For sequencing analysis, the concordance rate was 100 % on negative P-IHC and positive AP-IHC-stained sections, regardless of the subcellular localization of the reaction product. Mutations were detected in only 52 % of cases expressing nuclear/nuclear-cytoplasmic markers, regardless of the sequencing technology used (p = 0.0002). In conclusion, stained sections may be a valuable resource for FISH or sequencing analysis, but on cases expressing nuclear markers sequencing results need to be interpreted cautiously.
| Original language | English |
|---|---|
| Pages (from-to) | 1-11 |
| Number of pages | 11 |
| Journal | Virchows Archiv |
| DOIs | |
| Publication status | Accepted/In press - Jan 27 2016 |
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Keywords
- Direct sequencing
- Fish
- Immunohistochemistry
- Next generation sequencing
ASJC Scopus subject areas
- Pathology and Forensic Medicine
- Cell Biology
- Molecular Biology
Cite this
Doing more with less : fluorescence in situ hybridization and gene sequencing assays can be reliably performed on archival stained tumor tissue sections. / Pelosi, Giuseppe; Perrone, Federica; Tamborini, Elena; Fabbri, Alessandra; Testi, Maria Adele; Busico, Adele; Settanni, Giulio; Picciani, Benedetta; Bovio, Enrica; Sonzogni, Angelica; Valeri, Barbara; Garassino, Marina; de Braud, Filippo; Pastorino, Ugo.
In: Virchows Archiv, 27.01.2016, p. 1-11.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Doing more with less
T2 - fluorescence in situ hybridization and gene sequencing assays can be reliably performed on archival stained tumor tissue sections
AU - Pelosi, Giuseppe
AU - Perrone, Federica
AU - Tamborini, Elena
AU - Fabbri, Alessandra
AU - Testi, Maria Adele
AU - Busico, Adele
AU - Settanni, Giulio
AU - Picciani, Benedetta
AU - Bovio, Enrica
AU - Sonzogni, Angelica
AU - Valeri, Barbara
AU - Garassino, Marina
AU - de Braud, Filippo
AU - Pastorino, Ugo
PY - 2016/1/27
Y1 - 2016/1/27
N2 - Little is known about molecular testing on tumor tissue retrieved from stained sections, for which there may be a clinical need. We retrospectively analyzed 112 sections from 56 tumor patients using either fluorescence in situ hybridization (FISH) with different probes (19 sections from 17 patients) or Sanger or targeted next generation sequencing for detection of BRAF, EGFR, KRAS, C-KIT, and TP53 mutations (93 sections from 39 patients). Tumor tissue sections had been stained by hematoxylin and eosin (H&E) (42 sections) or by immunohistochemistry for cytoplasmic or nuclear/nuclear-cytoplasmic markers (70 sections) with a peroxidase (P-IHC, with 3,3′-diaminobenzidine as chromogen) or alkaline phosphatase label (AP-IHC, with Warp Red™ as chromogen). For FISH analysis, the concordance rate between the original diagnosis and that obtained on H&E- or P-IHC-stained tissue sections (AP-IHC was not on record for this set of patients) was 95 % (18 out of 19 tumor sections). Only one tumor sample, diffusely positive for MLH1, did not yield any nuclear hybridization signal. For sequencing analysis, the concordance rate was 100 % on negative P-IHC and positive AP-IHC-stained sections, regardless of the subcellular localization of the reaction product. Mutations were detected in only 52 % of cases expressing nuclear/nuclear-cytoplasmic markers, regardless of the sequencing technology used (p = 0.0002). In conclusion, stained sections may be a valuable resource for FISH or sequencing analysis, but on cases expressing nuclear markers sequencing results need to be interpreted cautiously.
AB - Little is known about molecular testing on tumor tissue retrieved from stained sections, for which there may be a clinical need. We retrospectively analyzed 112 sections from 56 tumor patients using either fluorescence in situ hybridization (FISH) with different probes (19 sections from 17 patients) or Sanger or targeted next generation sequencing for detection of BRAF, EGFR, KRAS, C-KIT, and TP53 mutations (93 sections from 39 patients). Tumor tissue sections had been stained by hematoxylin and eosin (H&E) (42 sections) or by immunohistochemistry for cytoplasmic or nuclear/nuclear-cytoplasmic markers (70 sections) with a peroxidase (P-IHC, with 3,3′-diaminobenzidine as chromogen) or alkaline phosphatase label (AP-IHC, with Warp Red™ as chromogen). For FISH analysis, the concordance rate between the original diagnosis and that obtained on H&E- or P-IHC-stained tissue sections (AP-IHC was not on record for this set of patients) was 95 % (18 out of 19 tumor sections). Only one tumor sample, diffusely positive for MLH1, did not yield any nuclear hybridization signal. For sequencing analysis, the concordance rate was 100 % on negative P-IHC and positive AP-IHC-stained sections, regardless of the subcellular localization of the reaction product. Mutations were detected in only 52 % of cases expressing nuclear/nuclear-cytoplasmic markers, regardless of the sequencing technology used (p = 0.0002). In conclusion, stained sections may be a valuable resource for FISH or sequencing analysis, but on cases expressing nuclear markers sequencing results need to be interpreted cautiously.
KW - Direct sequencing
KW - Fish
KW - Immunohistochemistry
KW - Next generation sequencing
UR - http://www.scopus.com/inward/record.url?scp=84955619448&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84955619448&partnerID=8YFLogxK
U2 - 10.1007/s00428-016-1906-0
DO - 10.1007/s00428-016-1906-0
M3 - Article
C2 - 26818831
AN - SCOPUS:84955619448
SP - 1
EP - 11
JO - Virchows Archiv - A Pathological Anatomy and Histopathology
JF - Virchows Archiv - A Pathological Anatomy and Histopathology
SN - 0945-6317
ER -