PRODUCT BULLETIN

GeoMx® Human Whole Transcriptome Atlas
The GeoMx Human Whole Transcriptome Atlas (WTA) empowers researchers to explore biology across a tissue. WTA is compatible with common
sample types such as formalin-fixed paraffin embedded (FFPE) or fresh frozen (FF) tissue and across all human tissues types. The human wholetranscriptome is measured in each region of interest (ROI) to uncover biological changes at specific locations in the tissue.

Product Highlights•Whole transcriptome coverage with probes specific toprotein coding mRNA sequences•

Superior sensitivity to detect 1000s of unique humangenes in <50 μm regions•

Robust performance across sample types including FFPE
with high concordance with RNA-seq and RNAscope™

•

Map single cell RNA-seq populations to their tissuelocation•

Integrated Illumina library prep protocols andstreamlined NGS analysis pipelines•

Flexibility to customize with up to 60 additional targets toassay non-coding RNA, synthetic DNA or exogenous genes•

Visualization and statistical analysis using GeoMx Data
Suite and bioinformatics toolset

Gene Density

Discover a new way to look at biology

Low

The GeoMx Whole Transcriptome Atlas delivers the maximum amountof sensitivity and confidence in each transcript through its unique probearchitecture. The WTA profiles over 18,000 protein-coding human genes
based on the human gene nomenclature committee (HUGO1) databasecross-referenced with available mRNA sequences in the National Center
for Biotechnology’s Information (NCBI) RefSeq database2 (Figure 1). Alongwith a high specificity for their target molecules, DSP RNA probes arescreened for a myriad of factors including melting temperature, GC content,
off target hits, and repetitive sequences. Each probe is assigned a unique
DSP barcode for downstream molecular counting on an Illumina sequencer.
The targeted nature of WTA avoids the many pitfalls of poly(A) pulldown
by delivering increased sensitivity even in heavily degraded tissues such
as FFPE. With GeoMx WTA, researchers can explore pathways across thewhole transcriptome in user defined regions of interest.

FOR RESEARCH USE ONLY. Not for use in diagnostic procedures.

1

23456789High

10 11 12 13 14 15 16 17 18 19 20 21 22 XYFigure 1: The Human WTA covers over 18,000 protein-coding genes
cross-referenced with the HUGO and NCBI RefSeq databases.

1GeoMx® Human Whole Transcriptome Atlas

PRODUCT BULLETIN

A

Explore the biology that matters

B
ROI size (μm)

1.00

Genes Detected above the Limit ofQuatification

50
18000

200
400

0.50

0.25

16000

Genes Detected

0.75
sensitivity

The GeoMx WTA offers high sensitivity and specificity withthousands of genes detectable across regions of interest (ROI)of various sizes. In a sensitivity analysis in FFPE with cell pellet
arrays of colorectal carcinoma (CRC) and non-small cell lungcancer (NSCLC), over 5000 genes were detected above the limitof quantification.

14000
12000
10000
8000
6000
4000
2000

0.00

00.00

0.25

0.50

0.75

1.00

0100

200300

400

500

600

700

Region of Interest Diameter (μm)

1 − specificity

Figure 2: A) Using RNA-seq transcripts per million as the ground truth for the
expression of each gene, the ROC curve shows high sensitivity and specificity. B)
GeoMx WTA detects a large number of genes across many regions of various sizes.

Spatial RNA profiling alignswith RNASeq and RNAscopeAB
WTA vs RNAscope1.00
DAUDI0.75

HUH7
HUT780.50

JURKAT

0.25

SUDHL1SUDHL40.00
50250360

ROI size (μm)

sensitivity

Traditional gene expression technologies are unable to capture
heterogeneity of the transcriptome with spatial context.
Bulk RNA sequencing and single cell RNAseq have delivered
interesting clinical and cell type signatures but localizingthese signatures is challenging with protein-based IHC. Other,more precise, technologies like fluorescently-labeled in situ
hybridization lack the plex to truly make new discoveries. WTA
delivers both spatial precision and high plex. Furthermore, WTA
compares favorably to RNA sequencing and RNAscope (Figure3) and is able to confidently detect transcripts in small regions.

correlation to RNAseq

WTA vs RNAseq
1.00

ROI size (μm)(# of nuclei)

0.75

50 (13.4)
80 (31.0)110 (54.3)
250 (258.5)
360 (525.8)

0.50
0.25
0.00
1−2 2−5 5−10 10−20 >20

number of transcripts per cell

Figure 3: A) GeoMx Human WTA performance compared to RNA-seq with
3 ROI sizes run on formalin-fixed paraffin embedded (FFPE) cell pellet arrays.
B) Comparing RNAscope and WTA counts illustrates the high sensitivity of WTA
even with small ROI sizes.

Map the location of your cell populations
Spatial analysis of a non-small cell carcinoma sample using the GeoMx SpatialDecon algorithm3 reveals cell type abundance in eachregion of interest (ROI) (Figure 4). Researchers can use the pre-defined cell definitions or their single cell RNAseq data to assess cell typeabundance throughout the tissue.
AB

Figure 4: A) Segmenting ROIs in PanCK+ andPanCK- regions enables the comparison of tumor
and tumor microenvironment segments. B) Using
the GeoMx Spatial Decon algorithm, researchers
can estimate the abundance of mixed cell types
within each ROI and segment. Legend to rightindicates cell type.

FOR RESEARCH USE ONLY. Not for use in diagnostic procedures.

2NanoString Technologies, Inc.

GeoMx® Human Whole Transcriptome Atlas

GeoMx® WTA Workflow
GeoMx RNA assays contain in situ hybridization (ISH) probes conjugated to unique DNA indexing-oligonucleotides (DSP barcodes) via a UVphotocleavable linker. After selecting regions of interest (ROIs) on the GeoMx DSP, the DSP barcodes are UV cleaved and collected (Figure 5).
During library preparation, the DSP barcodes are tagged with their ROI location, then sequenced on an Illumina sequencer. DNA oligonucleotide
sequences contain ROI indices mapping them back to their tissue location, an RNA target identification sequence matching them to their ISH
probes, and a unique molecular identifier (UMI) to deduplicate reads. Sequenced oligonucleotides are processed and then imported back into theGeoMx DSP analysis software for integration with the slide images and ROI selections for spatially-resolved RNA expression.

1

Stain slides with RNAscope™ probes
and GeoMx® DSP oligo-conjugated
RNA detection probes

RNAscope™
ProbesGeoMx®

2

Select Regions ofInterest (ROI)

3

UV-cleave and collect
DSP barcodes oﬀ RNA
probes in ROI

4

Dispense oligos
into 96-well plate

6

Construct
Library,Sequence& Count

DSP Barcoded
RNA probes

DigitalSpatial Profiler

DSP Barcoded
RNA Probes

DSP
BarcodedprobesFluorescentantibodies

5

Repeat for each ROI

Figure 5 : GeoMx DSP workflow enables the analysis of whole transcriptome data from user-defined regions.

A

GeoMx® Data Analysis

B
Glomerulus

DistalTubules
ProximalTubules
2 mm

100 µmCDE
Glomeruli

Proximal

DistalCell Abundance Estimate (%)

GeoMx software uniquely combines whole tissue visualization
at single cell resolution with advanced ROI selection to enable
comprehensive spatial profiling of tissue sections. The fully
integrated workflow tracks image data to corresponding profilingdata, allowing users to easily go from data collection to data
analysis and to interact with either data type in real time. The
data analysis suite provides multiple options to assess the qualityof the raw data and various methods to normalize data sets.
Moreover, multiple analysis methods are available, including
statistical methods, and cluster and pathway analysis. Multiple
data visualizations are available to enable data exploration andare then exportable as publication-quality figures. Visualization
plots include: heatmap, dendograms, bar graph, box plot, scatterplot, line/trend plot, volcano plot, forest plots, and 3-D PCA.
Analysis and visualization can be extended through the integrationof R-scripts into the Data Analysis Suite which are available on
GeoScript Hub, or through export of the raw or normalized data
to external tools including our GeoMxTools R package available in
Bioconductor™.

Figure 6: A-B) ROI selection strategy in kidney to study normal and diseased (DKD) kidney C) tSNE representation of glomerulus (magenta), proximal (green) anddistal (gray) tubules D) Differential expression showing genes from scRNA Seq expressed in podocytes (blue), proximal (green) or distal (red) tubules. E) Cell typeabundance estimates based on scRNA Seq from kidney samples.
FOR RESEARCH USE ONLY. Not for use in diagnostic procedures.

3PRODUCT BULLETIN

GeoMx® Human Whole Transcriptome Atlas

Ordering Information
GeoMx Whole Transcriptome Atlas
ProductProduct Description

GeoMx Human Whole Transcriptome Atlas
Human RNA for Illumina Systems

RNA Probe set that targets 18,000+ transcripts for human protein coding
genes plus ERCC negative controls to profile the whole transcriptome.
Excludes uninformative high expressing targets such as ribosomal
subunits. Includes RNA probes designed for Illumina NGS readout with
the Seq Code library prep.

Quantity Catalog Number

4 Slides

GMX-RNA-NGSHuWTA-4

GeoMx Mouse Whole Transcriptome Atlas
Mouse RNA for Illumina Systems

RNA Probe set that targets 21,000+ transcripts for mouse protein
coding genes plus ERCC negative controls to profile the wholetranscriptome. Excludes uninformative high expressing targets suchas ribosomal subunits. Includes RNA probes designed for Illumina NGS
readout with the Seq Code library prep.

4 Slides

GMX-RNA-NGSMsWTA-4

GeoMx Morphology Kits
ProductProduct Description

Quantity Catalog Number

GeoMx Solid Tumor TME Morphology Kit
Human RNA Compatible

Morphology kit for visualization of human solid tumors and the tumor
microenvironment. For use with RNA assays. Includes fluorescentantibodies against Pan-CK, CD45, and a nuclear stain.

12 slides

GMX-RNA-MORPHHST-12

GeoMx Melanoma TME
Morphology Kit
Human RNA Compatible

Morphology kit for visualization of human melanoma and the tumor
microenvironment. For use with RNA assays. Includes fluorescentantibodies against S100B/Pmel17, CD45, and a nuclear stain.

12 slides

GMX-RNA-MORPHHMEL-12

Additional Assay Reagents
ProductProduct Description

Quantity Catalog Number

GeoMx Seq Code Pack
Compatible with Illumina Systems

NGS readout reagents for GeoMx DSP RNA and protein analysis. Includes
two Seq Code primer plates (choice of A&B, C&D, E&F, or G&H) and two
universal enzyme master mixes.

192 AOI

GMX-NGS-SEQ-[XX]GeoMx RNA Slide Prep Kit for FFPE

Sample prep reagents for GeoMx DSP RNA analysis. Includes Buffer W,Buffer S, and Buffer R.

12 slides

GMX-PREP-RNAFFPE-12

GeoMx DSP Collection Plate

Barcoded collection plates for use on the GeoMx DSP. Required for AOI
tracking. Kit includes 12 plates
covering 1,152 AOI.

1 PackGMX-DSP-COLL-PLT

GeoMx DSP Instrument Buffer Kit

Buffer kit for the GeoMx DSP. Includes Buffer S and Buffer H. Sufficientfor ~48 samples with ~18 AOI each. Volume requirements may vary basedon experimental design.

1 Kit

GMX-DSP-BUFF-KIT

Selected Panel References
1.

HUGO Gene Nomenclature Committee. https://www.genenames.org/

2.

RefSeq: NCBI Reference Sequence Database. https://www.ncbi.nlm.nih.gov/refseq/

3.

Danaher, et al. Advances in mixed cell deconvolution enable quantification of cell types in spatially-resolved gene expression data. bioRxiv pre-print.

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FOR RESEARCH USE ONLY. Not for use in diagnostic procedures.
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