Journal: bioRxiv

Article Title: A new method for the sampling and preservation of placental specimens in low-resource settings for histology and analysis of nucleic acids

doi: 10.1101/2022.01.12.476059

Figure Lengend Snippet: RNAscope ® hybridization with probes for three genes. A . DAPI B . MEG3 C . ERVW-1 D . DIO3OS. E . Merged. Scale bar is 20 μm.

Article Snippet: RNAscope slides were imaged on a Zeiss Apotome microscope.

Techniques: Hybridization

Journal: Genome Biology

Article Title: Data-driven identification of total RNA expression genes for estimation of RNA abundance in heterogeneous cell types highlighted in brain tissue

doi: 10.1186/s13059-023-03066-w

Figure Lengend Snippet: Overview of the smFISH RNAscope experiment and DLPFC anatomy. a Illustration of RNAscope experimental design where a single DLPFC tissue block was used to generate 9 spatially adjacent slices. These 9 slices were hybridized with 3 RNAscope probe combinations noted as the AKT3 , ARID1B , and MALAT1 / POLR2A experiments (related to Additional file : Tables S4-S5). Candidate TREGs and POLR2A are shown in black, while GAD1 , SLC17A7 , and MBP are cell-type marker genes for inhibitory neurons (red), excitatory neurons (blue), and oligodendrocytes (orange), respectively. b Annotated image of DLPFC tissue, noting the location of gray matter (GM), white matter (WM), and sulcus. c Spatial distribution of cells expressing MBP for each sample. MBP is an oligodendrocyte cell type marker gene enriched in white matter

Article Snippet: Segmentation was optimized for each dye for each tissue section by adjusting several values with reference to the manufacturer’s guidelines: HALO 3.3 FISH-IF Step-by-Step guide (Indica Labs, version 2.1.4 July 2021) and Quantitative RNAscope Image Analysis Guide (Indica Labs).

Techniques: Blocking Assay, Marker, Expressing

Journal: Genome Biology

Article Title: Data-driven identification of total RNA expression genes for estimation of RNA abundance in heterogeneous cell types highlighted in brain tissue

doi: 10.1186/s13059-023-03066-w

Figure Lengend Snippet: Expression of TREGs in individual nuclei using smFISH with RNAscope. Representative high-magnification images showing the expression of candidate TREGs a AKT3 , b ARID1B , c HK gene POLR2A , and d MALAT1 and in human DLPFC. Insets show individual nuclei with high expression (yellow arrow), low expression (green arrow), or in rare cases (≤ 14% for candidate TREGs and 22% for POLR2A , Table ) no expression (purple arrow). Each puncta represents a single transcript, as illustrated in Fig. a. MALAT1 shows extremely high expression in the majority of nuclei such that individual puncta cannot be quantified (yellow arrow). Scale bar is 20 um

Article Snippet: Segmentation was optimized for each dye for each tissue section by adjusting several values with reference to the manufacturer’s guidelines: HALO 3.3 FISH-IF Step-by-Step guide (Indica Labs, version 2.1.4 July 2021) and Quantitative RNAscope Image Analysis Guide (Indica Labs).

Techniques: Expressing

Journal: Genome Biology

Article Title: Data-driven identification of total RNA expression genes for estimation of RNA abundance in heterogeneous cell types highlighted in brain tissue

doi: 10.1186/s13059-023-03066-w

Figure Lengend Snippet: Proportion of nuclei that displayed any TREG candidate or POLR2A puncta. Proportion of nuclei with a non-zero count in the DLPFC snRNA-seq data compared against the mean proportion of non-zero puncta in the nucleus and mean number of puncta observed in the RNAscope data for the candidate TREGs and POLR2A . Beta values are the slope of the linear fit of the number of puncta over ordered cell types and the 95% confidence interval. The standardized beta is the slope of the linear fit of the number of puncta divided by the standard deviation of the number of puncta for each gene. Standardized betas enable the comparison between snRNA-seq and RNAscope data. The standardized beta in snRNA-seq is − 1.33 (− 1.35, − 1.31). With RNAscope, AKT3 is the TREG that most similarly follows the trend across all genes in snRNA-seq (see also Fig. 7 ). Due to the inability to resolve individual puncta for MALAT1 , the observed trend (Additional file 2 : Fig. S10) is unreliable

Article Snippet: Segmentation was optimized for each dye for each tissue section by adjusting several values with reference to the manufacturer’s guidelines: HALO 3.3 FISH-IF Step-by-Step guide (Indica Labs, version 2.1.4 July 2021) and Quantitative RNAscope Image Analysis Guide (Indica Labs).

Techniques: Standard Deviation, Comparison

Journal: Genome Biology

Article Title: Data-driven identification of total RNA expression genes for estimation of RNA abundance in heterogeneous cell types highlighted in brain tissue

doi: 10.1186/s13059-023-03066-w

Figure Lengend Snippet: Boxplots of total RNA nuclear expression in the nucleus across cell types. a Distribution of total nuclear RNA expression (estimated with the sum of total UMIs per nucleus) in DLPFC snRNA-seq data across excitatory neurons (Excit), inhibitory neurons (Inhib), and oligodendrocytes (Oligo). b Distribution of the number of puncta quantified by RNAscope for each observed gene across the same cell types as in a . The number of puncta by RNAscope estimates the total RNA expression by snRNA-seq (Fig. a). POLR2A was only evaluated in excitatory neurons and oligodendrocytes as it was multiplexed with MALAT1 and GAD1 was omitted

Article Snippet: Segmentation was optimized for each dye for each tissue section by adjusting several values with reference to the manufacturer’s guidelines: HALO 3.3 FISH-IF Step-by-Step guide (Indica Labs, version 2.1.4 July 2021) and Quantitative RNAscope Image Analysis Guide (Indica Labs).

Techniques: Expressing, RNA Expression, Inhibition

Journal: Cancers

Article Title: Unique Spatial Immune Profiling in Pancreatic Ductal Adenocarcinoma with Enrichment of Exhausted and Senescent T Cells and Diffused CD47-SIRPα Expression

doi: 10.3390/cancers12071825

Figure Lengend Snippet: CD8+ T lymphocytes in the tumor center (TC) and the invasive front (IF) exhibit an exhausted phenotype. ( A ) RNAscope analysis to examine the co-expression of HAVCR2 (green) and/or PDCD1 (red) and CD8A (yellow) in PDAC patients. Representative confocal micrographs in PDAC patients without (w/o) neoadjuvant therapy and PDAC patients who received neoadjuvant chemotherapy. Dashed line delineates the invasive front (IF). Yellow asterisks depict cancer glands. Double arrowheads indicate CD8A -positive cells co-expressing HAVCR2/PDCD1 and single arrowheads depict only CD8A expressing T lymphocytes. Scale bar: 100 μm ( B ) Quantification of CD8+ T lymphocytes expressing HAVCR2/PDCD1 mRNA in PDAC patients who did not receive neoadjuvant therapy. ** p -value < 0.01. Data are expressed as mean ± STDEV (N = 5). ( C ) Quantification of CD8+ T lymphocytes expressing HAVCR2/PDCD1 mRNA in PDAC patients who received neoadjuvant chemotherapy. ** p -value < 0.01. Data are expressed as mean ± STDEV (N = 3). Tumor center (TC), invasive front (IF), normal parenchyma adjacent to the tumor (NAT).

Article Snippet: A restricted funding was received by GlaxoSmithKline to support RNAscope analysis.

Techniques: RNAscope, Expressing