Formalin-fixed, paraffin-embedded (FFPE) samples are valuable resources for translational medicine studies. Global gene expression profiling of FFPE material is challenging as the quality and availability of recovered RNA is heavily hampered. ArrayXS Human FFPE compensates for this deficit and enables reliable and robust detection of the expression of 39,975 protein coding genes in FFPE samples - even in tiny amounts as obtained from laser capture microdissection (LCM).
Perfect harmonisation of experimentally validated probes, RNA isolation and processing are key to high-quality data which you receive for every project. Combined with our dedicated sample processing protocol, ArrayXS Human FFPE delivers highest data quality for FFPE materials.
Sourced from Annotation Release 106 based on the genome assembly GRCh38; 33,975 genes are represented on ArrayXS Human FFPE.
Our turnaround times starting from 2 weeks including data analysis guarantee fast progress in your research project.
Agilent’s superior sensitivity, the extended dynamic range over 5 logs and OakLabs’ proven validation strategy allow to detect subtle biological changes with confidence. Our specialized lab team can even work with tiny amounts as obtained from laser capture microdissection (LCM).
ArrayXS Human FFPE is validated and certified for new strong features creating an outstanding experience for your R&D.
Bundled with a comprehensive data analysis package, the ArrayXS service enables a rapid interpretation of results.
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|OakLabs product number||XS-200110|
|GEO Platform ID||to be announced|
|Design creation date||12/20/2015|
|Format||8 x 60K|
|Biological features||33,975 target IDs (coding genes)|
|Replicates of biological probes||300 x 10|
|Positive controls||35 x 12 ERCC control oligonucleotides 35 x 10 E1A spike-in control oligonucleotides|
|Gene list, annotations, design files, probe sequences||Contact service(at)oak-labs.com.|
|Composition||Sequence content sourced from: genome build GRCh38, probe design and protocol adapted to FFPE samples|
|Validation details||OakLabs’ approved MyArray strategy (details: Development of ArrayXS Human FFPE)|
|Manufacturing||Agilent 60-mer SurePrint technology, details: Agilent’s Microarray Technology|
|Compatibility||1- and 2-colour hybridisations|
1. DEVELOPMENT OF AN INITIAL TEST ARRAY
An initial large microarray was developed to detect 33,975 coding transcripts. Every sequence was represented by up to 20 different probes. In addition, all probes of Agilent’s SurePrint G3 Human Gene Expression 8x60K v2 Microarray (ID 039464) as well as OakLabs ArrayXS Human (ID 079407) were added.
2. HYBRIDISATION OF VARIOUS SAMPLES
FFPE samples of various sources and ages were processed and hybridised onto the initial microarrays.
3. SELECTION OF THE BEST PROBES
Based on the hybridisation data, the best suited probes were selected to reliably measure gene expression in FFPE samples.
ARRAYXS HUMAN FFPE'S PROBES PERFORM BETTER
Each probe on ArrayXS Human FFPE was selected from a variety of different probes, including those of the 8x60K Agilent microarray and OakLabs' ArrayXS Human, both not developed specifically for FFPE samples. For the majority of genes, neither the Agilent nor the ArrayXS Human's probe was the best choice.
Figure 1 illustrates, exemplarily for one gene, the probes’ signals on ArrayXS Human FFPE (green shadowed) compared to other tested probes including the one on Agilent’s 8x60K (red shadowed) as well as on OakLabs ArrayXS Human (blue shadowed).
The importance of harmonisation of probe design and sample processing and hybridisation to achieve high data quality is also illustrated in figures 2 and 3. The histograms show the signal distributions of the same sample hybridised to ArrayXS Human FFPE (blue) compared to the alternative microarrays (grey), Agilent's 8x60K (Fig 2) and ArrayXS Human (Fig 3). In both comparisons, the histograms of the signals obtained from ArrayXS Human FFPE are shifted to the right towards larger signals and at the same time, the peak on the left is significantly smaller indicating that the expression of a large portion of genes can be detected only with ArrayXS Human FFPE. > .
2-COLOUR DYE SWAP APPROACH FOR STRINGENT AND CONFIDENT GENE IDENTIFICATION
In gene expression studies from fresh material, a 1-colour approach is preferred over a 2-colour approach due to its large flexibility and the lower costs.
In contrast, a 2-colour dye swap analysis of FFPE material is capable of increasing the confidence of the identified genes, as demonstrated in a pilot project that has been performed on tumor and normal tissue of 10 patients.
The mean log2 fold changes of tumor versus normal are visualised in a scatter plot (figure 4). The x-axis shows the data with the tumor tissue cy5-labeled and the normal tissue cy3-labelled, and y-axis vice versa.
Genes which have been identified to be significantly (at a p-value of 0.05) differentially expressed in both labelling orientations appear green. Those ones that are not significantly differentially expressed in either orientation appear in the middle of the plot and grey.
Those genes with detection of signficant differential expression in only one labelling orientation appear red. Those genes illustrate the portion of potentially false positive identifications if a dyes swap approach is being avoided. Therefore, by a 2-colour dye swap approach the significance of the identified genes will be increased.