Combined single-molecule fluorescence in situ hybridization and immunohistochemistry analysis in intact murine dorsal root ganglia and sciatic nerve

STAR Protocols

Mollie K. Meffert¹  Sreenivas Eadara²  Xinbei Li²  Michael J. Caterina²  Yan Liu³  Gabriella Muwanga³  Lintao Qu³  Sangmin Jeon³ 

[1] Department of Neurosurgery and Neurosurgery Pain Research Institute, Baltimore, MD 21205, USA;Department of Biological Chemistry, The Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA;Department of Neurosurgery and Neurosurgery Pain Research Institute, Baltimore, MD 21205, USA;

关键词: Microscopy;    Molecular Biology;    In Situ Hybridization;    Neuroscience;    Molecular/Chemical Probes;   

DOI  :  

来源: DOAJ

【 摘 要 】

Summary: Single-molecule fluorescence in situ hybridization (smFISH) allows spatial mapping of gene expression. This protocol presents advances in smFISH fidelity and flexibility in intact murine sensory nervous system tissue. An approach using RNAscope probes allows multiplexing, enhanced target specificity, and immunohistochemistry compatibility. Computational strategies increase quantification accuracy of mRNA puncta with a point spread function for clustered transcripts in the dorsal root ganglion and 3D masking for intermingled sciatic nerve cell types. Approaches are validated for mRNAs of modest (Lin28a) and medium (Ppib) steady-state abundance in neurons.

【 授权许可】

Unknown