Preparing Planarian Cells for High-Content Fluorescence Microscopy Using RNA in Situ Hybridization and Immunocytochemistry.
Methods Mol Biol
; 2680: 121-155, 2023.
Article
en En
| MEDLINE
| ID: mdl-37428375
High-content fluorescence microscopy combines the efficiency of high-throughput techniques with the ability to extract quantitative information from biological systems. Here we describe a modular collection of assays adapted for fixed planarian cells that enable multiplexed measurements of biomarkers in microwell plates. These include protocols for RNA fluorescent in situ hybridization (RNA FISH) as well as immunocytochemical protocols for quantifying proliferating cells targeting phosphorylated histone H3 as well as 5-bromo-2'-deoxyuridine (BrdU) incorporated into the nuclear DNA. The assays are compatible with planarians of virtually any size, as the tissue is disaggregated into a single-cell suspension before fixation and staining. By sharing many reagents with established planarian whole-mount staining protocols, preparation of samples for high-content microscopy adoption requires little additional investment.
Palabras clave
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Planarias
/
ARN
Límite:
Animals
Idioma:
En
Revista:
Methods Mol Biol
Asunto de la revista:
BIOLOGIA MOLECULAR
Año:
2023
Tipo del documento:
Article
País de afiliación:
Alemania
Pais de publicación:
Estados Unidos