Evaluation of the Suitability of RNAscope as a Technique to Measure Gene Expression in Clinical Diagnostics: A Systematic Review

Abstract

Objective To evaluate the application of RNAscope in the clinical diagnostic field compared to the current ‘gold standard’ methods employed for testing gene expression levels, including immunohistochemistry (IHC), quantitative real time PCR (qPCR), and quantitative reverse transcriptase PCR (qRT-PCR), and to detect genes, including DNA in situ hybridisation (DNA ISH). Methods This systematic review searched CINAHL, Medline, Embase and Web of Science databases for studies that were conducted after 2012 and that compared RNAscope with one or more of the ‘gold standard’ techniques in human samples. QUADAS-2 test was used for the evaluation of the articles’ risk of bias. The results were reviewed narratively and analysed qualitatively. Results A total of 27 articles (all retrospective studies) were obtained and reviewed. The 27 articles showed a range of low to middle risk of bias scores, as assessed by QUADAS-2 test. 26 articles studied RNAscope within cancer samples. RNAscope was compared to different techniques throughout the included studies (IHC, qPCR, qRT-PCR and DNA ISH). The results confirmed that RNAscope is a highly sensitive and specific method that has a high concordance rate (CR) with qPCR, qRT-PCR, and DNA ISH (81.8–100%). However, the CR with IHC was lower than expected (58.7–95.3%), which is mostly due to the different products that each technique measures (RNA vs. protein). Discussion This is the first systematic review to be conducted on the use of RNAscope in the clinical diagnostic field. RNAscope was found to be a reliable and robust method that could complement gold standard techniques currently used in clinical diagnostics to measure gene expression levels or for gene detection. However, there were not enough data to suggest that RNAscope could stand alone in the clinical diagnostic setting, indicating further prospective studies to validate diagnostic accuracy values, in keeping with relevant regulations, followed by cost evaluation are required. Supplementary Information The online version contains supplementary material available at 10.1007/s40291-021-00570-2.

Spatially Resolved and Highly Complexed Protein and RNA in situ Detection by Combining CODEX with RNAscope In Situ Hybridization

“…CODEX represents a powerful new platform for highly multiplexed proteins (up to 60) in situ detection (6, 8, 20). RNAscope represents another power multiplex RNA detection system (up to 12) at a single-copy sensitivity and high specificity (19, 23), where the unique double Z probes and corresponding amplification probes enables concurrent signal amplification and background noise reduction. The combination of RNAscope with regular immunohistochemical or immunofluorescent staining to concurrently detect RNA and protein within the same tissue section has been reported by us (29) and other (30, 31).…”

“…Simultaneous visualization of spatially resolved and highly multiplexed proteins and RNA in situ is highly desirable for both basic and applied biomedical research (21, 22). RNAscope in situ hybridization (RNAscope) from Advanced Cell Diagnostics (ACD) is a novel RNA detection system with single-copy sensitivity and unprecedent specificity at a single cell level (19, 23). Nevertheless, RNAscope and CODEX involve complex chemical and physical procedures.…”

“…RNAscope in situ hybridization (RNAscope) from Advanced Cell Diagnostics (ACD) is a novel RNA detection system with single-copy sensitivity and unprecedent specificity at a single cell level (19,23). Nevertheless, RNAscope and CODEX involve complex chemical and physical procedures.…”

Draft Genome Assembly and Annotation for Cutaneotrichosporon dermatis NICC30027, an Oleaginous Yeast Capable of Simultaneous Glucose and Xylose Assimilation

“…Numerous methods for measuring the quantities of mRNA molecules and proteins can be utilized to determine the expression level of anticipated genes [ 65 ]. Reverse transcription-polymerase chain reaction (RT-PCR), northern blotting, microarrays, and transcriptome sequencing are all common methods for determining mRNA levels [ 66 ]. Protein levels can be determined using enzyme-linked immunosorbent assays (ELISA) and western blotting, as well as protein microarrays and proteomic analysis [ 67 , 68 ].…”

Spatially Resolved and Highly Multiplexed Protein and RNA In Situ Detection by Combining CODEX With RNAscope In Situ Hybridization

“…21,22 RNAscope in situ hybridization (RNAscope) from the Advanced Cell Diagnostics (ACD) is a novel RNA detection system with high sensitivity and unprecedent specificity at a single-cell level. 19,23 Nevertheless, RNAscope and CODEX involve complex chemical and physical procedures. To our knowledge, the combination of CODEX and RNAscope (Comb-CODEX-RNAscope) remained unknown until this study.…”