Tissue-Specific RNA Localization in the Uterus During Implantation, Decidualization and Placentation: Technical Nuances of Various Labeling Approaches.
FISH
RNA probe
digoxigenin
in situ hybridization
radioactive
tyramide signal amplification
Journal
Current protocols
ISSN: 2691-1299
Titre abrégé: Curr Protoc
Pays: United States
ID NLM: 101773894
Informations de publication
Date de publication:
Jun 2023
Jun 2023
Historique:
pmc-release:
01
06
2024
medline:
26
6
2023
pubmed:
22
6
2023
entrez:
22
6
2023
Statut:
ppublish
Résumé
In situ hybridization (ISH) is a sensitive method used to localize a specific sequence of DNA or RNA in biological samples, including cells, tissue sections or whole organs. RNA ISH can be used to determine spatial gene expression using a single-stranded probe with a reverse-complementary sequence. Cell-specific gene expression has been studied using mRNA and protein levels. Signals produced by RNA probes are usually more specific than those produced by antibodies in immunostaining. Currently, ISH is the most widely used method to localize mRNA molecules. Traditionally, probes were labeled with radioactive isotopes, but the cumbersome procedures and potential health risk limit their acceptance. Recently, probes labeled with nonradioactive materials including digoxigenin, biotin and various fluorophores have been developed. The tyramide signal amplification system further enhances the sensitivity of detection. These methods have been applied in numerous studies in various tissues including reproductive organs. This article details three methods of RNA in situ hybridization: radioactive in situ hybridization, digoxigenin in situ hybridization, and digoxigenin-tyramide signal amplification fluorescein in situ hybridization. The pros and cons of each protocol are discussed. © 2023 Wiley Periodicals LLC. Basic Protocol 1: Radioactive in situ hybridization (radioactive-ISH) Basic Protocol 2: Digoxigenin in situ hybridization (DIG-ISH) Basic Protocol 3: Digoxigenin-tyramide signal amplification fluorescein in situ hybridization (DIG-TSA-FISH).
Identifiants
pubmed: 37345986
doi: 10.1002/cpz1.823
pmc: PMC10299818
mid: NIHMS1906369
doi:
Substances chimiques
Digoxigenin
NQ1SX9LNAU
RNA, Messenger
0
RNA
63231-63-0
Fluoresceins
0
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
e823Subventions
Organisme : NICHD NIH HHS
ID : R01 HD068524
Pays : United States
Organisme : NICHD NIH HHS
ID : R01 HD103475
Pays : United States
Informations de copyright
© 2023 Wiley Periodicals LLC.
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