A modified immunofluorescence in situ hybridization method to detect long non-coding RNAs and proteins in frozen spinal cord sections.

Abstract

Immunofluorescence in situ hybridization (immuno-FISH) is widely used to co-detect RNAs and proteins in order to study their spatial distribution in cells. The present study used a modified immuno-FISH protocol for the detection of long non-coding RNAs (lncRNAs) and proteins in frozen spinal cord sections. The spinal cords of Sprague-Dawley rats were harvested, frozen and sectioned (10 µm), and oligonucleotide probes and antibodies were prepared. Following antigen retrieval, dehydration, prehybridization, hybridization, post-hybridization and immunofluorescence staining, images were captured. Antigen retrieval was performed by autoclaving or proteinase K treatment, and their effects on the hybridization signal were compared. The same sections were successfully stained by immunofluorescence. Satisfactory fluorescent signals of lncRNA and protein were obtained. The results of the present study suggest that the modified protocol of immuno-FISH for the detection of lncRNAs and proteins in frozen spinal cord sections is effective and time-efficient, and the required reagents are readily available.