title: Fully Automated RNAscope In Situ Hybridization Assays for Formalin-Fixed Paraffin-Embedded Cells and Tissues.
abstract: Biomarkers such as DNA, RNA, and protein are powerful tools in clinical diagnostics and therapeutic development for many diseases. Identifying RNA expression at the single cell level within the morphological context by RNA in situ hybridization provides a great deal of information on gene expression changes over conventional techniques that analyze bulk tissue, yet widespread use of this technique in the clinical setting has been hampered by the dearth of automated RNA ISH assays. Here we present an automated version of the RNA ISH technology RNAscope that is adaptable to multiple automation platforms. The automated RNAscope assay yields a high signal-to-noise ratio with little to no background staining and results comparable to the manual assay. In addition, the automated duplex RNAscope assay was able to detect two biomarkers simultaneously. Lastly, assay consistency and reproducibility were confirmed by quantification of TATA-box binding protein (TBP) mRNA signals across multiple lots and multiple experiments. Taken together, the data presented in this study demonstrate that the automated RNAscope technology is a high performance RNA ISH assay with broad applicability in biomarker research and diagnostic assay development. J. Cell. Biochem. 117: 2201-2208, 2016. © 2016 Wiley Periodicals, Inc.
authors.affiliation: Advanced Cell Diagnostics, 7707 Gateway Blvd, Newark, California, 94560.
authors.forename: Courtney M
authors.initials: CM
authors.lastname: Anderson
authors.affiliation: Advanced Cell Diagnostics, 7707 Gateway Blvd, Newark, California, 94560.
authors.forename: Bingqing
authors.initials: B
authors.lastname: Zhang
authors.affiliation: Advanced Cell Diagnostics, 7707 Gateway Blvd, Newark, California, 94560.
authors.forename: Melanie
authors.initials: M
authors.lastname: Miller
authors.affiliation: Advanced Cell Diagnostics, 7707 Gateway Blvd, Newark, California, 94560.
authors.forename: Emerald
authors.initials: E
authors.lastname: Butko
authors.affiliation: Advanced Cell Diagnostics, 7707 Gateway Blvd, Newark, California, 94560.
authors.forename: Xingyong
authors.initials: X
authors.lastname: Wu
authors.affiliation: Advanced Cell Diagnostics, 7707 Gateway Blvd, Newark, California, 94560.
authors.forename: Thomas
authors.initials: T
authors.lastname: Laver
authors.affiliation: Advanced Cell Diagnostics, 7707 Gateway Blvd, Newark, California, 94560.
authors.forename: Casey
authors.initials: C
authors.lastname: Kernag
authors.affiliation: Advanced Cell Diagnostics, 7707 Gateway Blvd, Newark, California, 94560.
authors.forename: Jeffrey
authors.initials: J
authors.lastname: Kim
authors.affiliation: Advanced Cell Diagnostics, 7707 Gateway Blvd, Newark, California, 94560.
authors.forename: Yuling
authors.initials: Y
authors.lastname: Luo
authors.affiliation: Advanced Cell Diagnostics, 7707 Gateway Blvd, Newark, California, 94560.
authors.forename: Henry
authors.initials: H
authors.lastname: Lamparski
authors.affiliation: Advanced Cell Diagnostics, 7707 Gateway Blvd, Newark, California, 94560.
authors.forename: Emily
authors.initials: E
authors.lastname: Park
authors.affiliation: Advanced Cell Diagnostics, 7707 Gateway Blvd, Newark, California, 94560.
authors.forename: Nan
authors.initials: N
authors.lastname: Su
authors.affiliation: Advanced Cell Diagnostics, 7707 Gateway Blvd, Newark, California, 94560.
authors.forename: Xiao-Jun
authors.initials: XJ
authors.lastname: Ma
mesh_terms: D001331:Automation; D014408:Biomarkers, Tumor / Q000032:analysis / Q000235:genetics*; D005404:Fixatives / Q000737:chemistry*; D005557:Formaldehyde / Q000737:chemistry*; D006367:HeLa Cells; D006801:Humans; D017403:In Situ Hybridization; D009369:Neoplasms / Q000235:genetics* / Q000473:pathology; D016612:Paraffin Embedding / Q000379:methods*; D012313:RNA / Q000235:genetics / Q000378:metabolism*
chemical_list: D014408:Biomarkers, Tumor; D005404:Fixatives; D005557:Formaldehyde; D012313:RNA
keywords: AUTOMATION; BIOMARKER; GENE EXPRESSION; IN SITU HYBRIDIZATION; RNA; RNAscope