Effect of formalin-fixation and paraffin-embedded tissue storage times on RNAscope in situ hybridization signal amplification.
J Vet Diagn Invest
; 36(4): 498-505, 2024 Jul.
Article
em En
| MEDLINE
| ID: mdl-38650110
ABSTRACT
RNAscope in situ hybridization (ISH) detects target RNA in formalin-fixed, paraffin-embedded (FFPE) tissues. Protocols suggest that prolonged FFPE storage and formalin fixation may impact signal detection, potentially limiting the utility of RNAscope ISH in retrospective studies. To develop parameters for RNAscope use with archived specimens, we evaluated the effect of formalin-fixation time by measuring the signal of a reference gene (16srRNA) in selected tissues fixed in 10% neutral-buffered formalin for 1, 2, 3, 5, 7, 10, 14, 21, 28, 60, 90, 180, and 270 d. The signal intensity and percent area of signal decreased after 180 d. Tissues had detectable signal at 180 d but not at 270 d of formalin fixation. To assess target detection in paraffin blocks, we qualitatively compared the signal of canine distemper virus (CDV) antigen via immunohistochemistry and CDV RNA via RNAscope ISH in replicate sections from blocks stored at room temperature for 6 mo, 1, 3, 6, 8, 11, 13, and 15 y; RNA was detected in FFPE tissues stored for up to 15 y. Our results demonstrate that RNAscope ISH can detect targets in tissues with prolonged paraffin storage intervals and formalin-fixation times.
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Texto completo:
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Base de dados:
MEDLINE
Assunto principal:
Fixação de Tecidos
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Inclusão em Parafina
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Hibridização In Situ
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Formaldeído
Limite:
Animals
Idioma:
En
Revista:
J Vet Diagn Invest
Assunto da revista:
MEDICINA VETERINARIA
Ano de publicação:
2024
Tipo de documento:
Article
País de afiliação:
Estados Unidos