Novel and simple method of double‑detection using fluorescence in�situ hybridization and fluorescence immunostaining of formalin‑fixed paraffin‑embedded tissue sections

Abstract

Abstract. The importance of fluorescence in situ hybridization (FISH) for pathological diagnosis has been increasing. However, the procedures utilized for a conventional FISH method with formalin-fixed paraffin-embedded tissue sections are complicated and it is difficult to perform as a routine laboratory test. In addition, there are difficulties with differentiation of targeted cells in observations with a fluorescence microscope. The present study reported a novel method that utilizes FISH in combination with fluorescence immunostaining as a simple double-detection technique that addresses these problems. Using this novel method, various genetic aberrations, as well as protein overexpression were easily visualized in isologous sections. In particular, FISH signals with our method clearly identify target cells in samples with poor differentiation between tumor cells coexisting with normal cells. It is proposed that this simple technique is widely applicable as a routine laboratory test and future developments are expected.

Ultra High-Plex Spatial Proteogenomic Investigation of Giant Cell Glioblastoma Multiforme Immune Infiltrates Reveals Distinct Protein and RNA Expression Profiles

“…These novel multimodal omic datasets of protein and DNA or RNA have been termed "spatial proteogenomics". However, until recently, spatial proteogenomic protocols with ultrahighplex, simultaneous co-detection of analytes have not been available (8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20)(21). Many of the protocols combine high-plex RNA with up to 4 fluorescent antibodies utilized as morphology markers.…”

Maintenance of Fluorescence During Paraffin Embedding of Fluorescent Protein-Labeled Specimens

“…For example, transgene and virus tracing with GFP-based reporter proteins provides a convenient way to explore neuronal morphology (Li et al, 2017, 2018), disease progression (Kammertoens et al, 2017), and the structure and function of neural circuits (Kim et al, 2015; Yao et al, 2018; Zhou et al, 2018). In order to combine paraffin embedding with fluorescent labeling techniques, antibodies are employed to detect GFP proteins as a compromise (Vankelecom, 2009; Porter et al, 2017; Ikeda, 2018). However, this compromising staining method is tedious and may result in either false negatives or false positives (True, 2008).…”