the agilent gene expression platform agile… · g4845a human ge 4x44k v2 microarray kit 4 x 44k 4...

The Agilent Gene Expression Platform

LINC-ing To The Future with Increased Density

Gene expression continues to be a core component of systems biology studies. As the

perspective of systems biology continues to change, it is imperative that the platforms involved

in gene expression continue to evolve. With that concept in mind, Agilent Technologies has

launched the new SurePrint G3 Gene Expression Microarray platform. The G3 GE platform has

undergone a redesign based on updated databases to ensure comprehensive coverage of

current content. In addition, the G3 GE platform has increased density to allow for expanded

content. One of the most exciting ways this expanded content is being utilized is with the

addition of non-coding RNAs, specifically lincRNAs, now making it possible to perform two

experiments from a single data set. Come see how this new design with updated and expanded

content can take your research into the future.

David Weiss

Field Application Scientist

Agilent Technologies

November 18, 2010

Systems BiologyKeys To A Successful Experiment

• Flexibility

• Sensitivity

• Dynamic Range

• Data Quality and Reproducibility

Agilent Confidential

For research use only. Not for use in

diagnostic procedures

Agilent Delivers With

A Complete Workflow!

FlexibilityCatalog and Custom Arrays through eArray• Wide range of catalog microarrays available

– Human, Mouse, Rat, Bovine, C.elegans, Canine, Chicken, Drosophila, Horse, Mosquito, Porcine,

Rabbit, Rhesus Macaque, Salmon, Sheep, Xenopus, Zebrafish

– Arabidopsis, Barley, Brassica, Cotton, Maize, Medicago, Rice, Tobacco, Tomato, Wheat

– E. coli, Magnaporthe, Yeast

• Unlimited custom microarray options in eArray

– Choose Agilent-optimized probes, upload your own sequences, or design probes

– Search and download Agilent catalog microarray content and include in your designs

ORANGE: Catalog arrays available in SurePrint G3 8x60K format

FlexibilityAgilent Microarray Manufacturing

AGILENT

- Industrial

manufacturing facility –

Class 10,000 clean-

room

- Ability to easily scale

production capacity –

NO delays

Nature Biotech. 19(4), 342-347 (2001).

FlexibilityG3 Format Features

• Comprehensive coverage

– Updated content

– Inclusion of non-coding RNAs, for the first time!

• Higher throughput for whole-genome studies

• Lower RNA input requirement (down to 10 ng)

• Increased affordability

• Increased density

– Combine species on a single array; for example, human + pathogens

– Target exons or splice junctions

– Replicate probes




SurePrint G3 Arrays

FlexibilityMultiple Gene Expression Microarray Configurations

SurePrint HD Arrays

1x

2x

4x

8x

65 µm features

Compatible with:

Agilent B or C Scanner

30 µm features

Compatible with:

Agilent C Scanner



60K15K

44K180K44K44K

44K400K44K105K

44K 1M44K 244K

SensitivityAccurate Representation of Low & High Abundance Transcripts

• All Agilent microarrays include probes for spike-in controls, which indicate labeling

and hybridization success

• Agilent spike-ins demonstrate

• Sensitive detection on the order of 1 in 6.9 million transcripts

• Detection across a dynamic range spanning 5 orders of magnitude

Dynamic RangeWide Dynamic Range for Sensitivity to Low Expressors

• Agilent gene expression data provides the expected normal distribution of signals, with few

probes in the noise

• Competing platforms suffer from compressed signals without a normal distribution

Dynamic RangeAdvantages

• 5 log Dynamic

Range allow detect

both high and low

abundant

transcripts

MCF-7

GO analysis for each Expression range

Metabolism

Biosynthesis

Regulation of transcription

Signal transduction

Data Quality & ReproducibilityComparison of Tumor & Normal Samples

• Various tissue samples and types provide similar levels of high

reproducibility using SurePrint G3 Gene Expression Microarrays

• Reproducibility translates in the ability to confidently identify genes of

differential expression



~5 o

rders

of m

agnitude

AdenocarcinomaNormal Colon

Replicate #2

Replicate #3

Rep

lic

ate

#1

Re

pli

ca

te #

2

Replicate #2

Rep

lic

ate

#

1

Replicate #3

Rep

lic

ate

#

2

Data Quality & ReproducibilityHigh Concordance and Correlation to TaqMan Data

• As compared with other microarrays providers, Agilent array gene expression signals span

a wider dynamic range

• With wide dynamic range, Agilent gene expression data reflect real gene expression levels

TaqMan Log Ratio

Mic

roa

rray L

og

Ra

tio

10 ng Input: n = 686, r = 0.93, m = 0.86

50 ng Input: n = 704, r = 0.94, m = 0.90

Agilent Array Correlation to Taqman



Data Quality & ReproducibilityIntra-Array Reproducibility

• High intra-array reproducibility indicated by low % coefficient of variance

between replicate probes on individual arrays

• SurePrint G3 arrays provide data you can trust

% C

V P

roce

sse

d S

ign

al

1x1M 2x400K 4x180K 8x60K

6

4

10

8

MAQC A

MAQC B



Data Quality & ReproducibilityInter-Array Reproducibility• High inter-array reproducibility indicated by low % coefficient of variance

between replicate probes on individual arrays

• SurePrint G3 arrays provide data you can trust

# G

en

es

De

tec

ted

CV

(%

)



Data Quality & ReproducibilitySurePrint G3 Microarrays and RNA-Seq: a Comparison

• Strong correlation of log ratios are observed

between microarray and RNA-Seq for

samples MAQC A and B

• Microarrays are faster, easier, more

affordable, and require little RNA input

Scatter Plot

B

M = 1.120

R = 0.840

N = 15516

RN

A-S

eq,

1 µ

g

8x60K, 25 ng, 1-color

Array and RNA-Seq

Log2 Ratios (B/A)

Agilent G3

+LIQARNA-Seq

Starting Material 10 ng 1 µg

Total Sample Prep 6.5 hr 11.5 hr

Hands-on Sample

Prep1.5 hr 3.75 hr

Sample to Data 1.5 days ~ 1 week

Analysis Simple Challenging

Price per sample $$ $$$



A Complete Solution for Gene Expression• RNA purification

• RNA quality assessment

• Catalog arrays for 30 species

• Unlimited custom array

options

• Simple assay with low RNA

input requirement

• Quality controls

• Biological interpretation

• qRT-PCR validation

Array Selection & Creation

Sample Labeling

Genome-wide & Custom ArraysHyb & Wash

Scanning and Feature Extraction

Data Analysis

Sample Prep and QA

qRT-PCR Validation

SurePrint G3

Exon Arrays

GeneSpring 11.5

Gene Expression Equipment List




Part Number Description Format

# Arrays per

Slide Qty/Kit Part Number Description Qty/Kit

G4851A SurePrint G3 Human GE 8x60K Kit 8x60K 8 3 5188-5279 Two-color RNA Spike-in Kit

G4852A SurePrint G3 Mouse GE 8x60K Kit 8x60K 8 3 5188-5282 One-color RNA Spike-in Kit

G4853A SurePrint G3 Rat GE 8x60K Kit 8x60K 8 3 5190-2305

Low Input Quick Amp Labeling

Kit, One-Color 24 rxns

G4102A SurePrint G3 Custom GE 8x60K 8x60K 8 1 5190-2306

Low Input Quick Amp Labeling

Kit, Two-Color 48 rxns

G4862A SurePrint G3 Custom GE 4x180K 4x180K 4 1 5188-5242

Gene Expression Hybridization

Kit

G4861A SurePrint G3 Custom GE 2x400K 2x400K 2 1 5185-5974 25X Fragmentation Buffer 10mls

G4860A SurePrint G3 Custom GE 1x1M 1x1M 1 1 5188-5281

10X Gene Expression Blocking

Agent, Large Volume

G4845A Human GE 4x44K v2 Microarray Kit 4 x 44K 4 5 5190-0403 2X Hi-RPM Hybridization Buffer 25mls

G4846A Mouse GE 4x44K v2 Microarray Kit 4 x 44K 4 5 5188-5325 Gene Expression Wash Buffer 1 4L

G4131F Whole Rat Genome Microarray Kit 4 x 44K 4 5 5188-5326 Gene Expression Wash Buffer 2 4L

G2514F Custom Microarray GE 4 x 44K 4 1 G2565CA

Agilent High-Resolution

Microarray Scanner Bundle

G4503A Custom Microarray GE 2 x 105K 2 1 G2534A

Hybridization Chamber,

Stainless

G4502A Custom Microarray GE 1 x 244K 1 1 G2534-60003 Backing Slides 5

G2509F Custom Microarray GE 8 x 15K 8 1 G2545A Hybridization Oven

G2530-60029 Hybridization Oven Rotator

G2939AA

2100 electrophoresis

Bioanalyzer (no PC!)

5067-1511 Agilent RNA 6000 Nano Kit 300

the agilent gene expression platform agile… · g4845a human ge 4x44k v2 microarray kit 4 x 44k 4...

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