GeneChip™ WT Pico Kit, 12 reactions - FAQs

Please refer to the Microarray Reagent Guide for Arrays and Expression Kits to match the correct reagents your array.

Find additional tips, troubleshooting help, and resources within our Microarray Analysis Support Center.

Sequences of all our controls can be downloaded from the sequence file that is available on product pages on thermofisher.com.

For example, here is the link to the product page for Clariom D Assay, human: https://www.thermofisher.com/order/catalog/product/902923
Go to the "Software & Data Analysis" section and under "Support Files", click on "Sequence Files: Clariom D Array, human Probe Sequences, tabular format", which you can open in Excel.

Note: If the control sequence file is not available, please contact techsupport@thermofisher.com.

Pseudogene databases were not included in the design of expression arrays.

The first dilution of the Poly-A RNA Control can be stored at -20 degrees C for 6 weeks (with up to 4 freeze/thaws).

Labeled material can be stored for 2 weeks at -20 degrees C.

Typical shelf life for expression arrays are as follows:
- All ≥ 8 µm commercial expression cartridges are stable up to 24 months.
- All ≥ 8 µm commercial resequencing cartridges are stable up to 18 months.
- All ≥ 8 µm commercial expression plates are stable up to 18 months.

Please note that this excludes products such as Exon or Gene array as they are 5 µm cartridge and plates.

For IVT assays, the RIN should be 6 or higher. For WT assays, there are no recommendations based on assay priming.

Find additional tips, troubleshooting help, and resources within our Microarray Analysis Support Center.

The GeneChip WT Pico kit is a strand-specific reagent, which must be used with Clariom D (and HTA 2.0) to obtain strand-specific information as these arrays having probes on regions overlapping from both strands. Strand-specificity is important for identifying antisense transcripts, determining the transcribed strand of ncRNAs, and demarcating the boundaries of closely situated or overlapping genes.
Strand-specificity is less important for gene-level expression analysis (i.e., when using Clariom S). Probes within regions of overlapping transcription from both strands are avoided in the Clariom S array design; i.e., the array is "stranded" meaning it does not necessarily need a strand-specific reagent kit.

Find additional tips, troubleshooting help, and resources within our Microarray Analysis Support Center.

The protocols for the Gene 1.0 ST and Exon 1.0 ST arrays are the same until the fragmentation and labeling steps.
Depending on the input amount, please refer to the GeneChip WT Pico Reagent Kit or the GeneChip WT PLUS Reagent Kit for the protocol:
- WT Pico: 50 - 500ng of total RNA
- WT PLUS: ≥ 100pg of total RNA (approx. 10 cells)
From the hybridization step onwards, the differences in protocol are due to the format of the arrays. The Exon array is a 49 format array, whereas the Gene 1.0 ST array is a 169 format array.
For the fluidics step the following scripts should be used depending upon which array you are using:
- Gene 1.0 St array: FS450_0001
- Exon 1.0 St array: FS450_0007

Find additional tips, troubleshooting help, and resources within our Microarray Analysis Support Center.

The GeneChip WT Pico Kit comprises reagents and a protocol for producing hybridization-ready DNA. The input for this kit is either from 100 pg to 10 ng (>500 pg recommended) of purified total RNA from cells or tissues, or 500 pg to 50 ng (>2 ng recommended) of purified total RNA from FFPE tissues.

Find additional tips, troubleshooting help, and resources within our Microarray Analysis Support Center.

Different methods are available to determine RNA concentration:
- UV Absorbance using a NanoDrop fluorospectrophotometer (5 to 500 ng/µL)
- Fluorescence-based quantitation (e.g. RiboGreen/NanoDrop Fluorospectrometer/ Qubit RNA HS Assay Kit and Qubit). Please check manufacturer recommendations for detection limits
- Bioanalyzer; the RNA Nano kit can be used to determine concentrations between 25 and 500 ng/µL. The RNA Pico kit should not be used for quantitation of RNA
- qRT-PCR; samples with concentrations below the detection limits of the afore mentioned methods may be quantified using qRT-PCR

Find additional tips, troubleshooting help, and resources within our Microarray Analysis Support Center.

We recommend using PAXgene tubes for collection or traditional collection and coagulation methods such as EDTA, heparin, etc., because PAXgene tubes will "lock" the blood profile.
For other methods, you would need to put in a preservative like TRIzol reagent, or to freeze the blood. After that, any RNA extraction/purification kit will work.

Find additional tips, troubleshooting help, and resources within our Microarray Analysis Support Center.

Please follow the instructions in the GeneChip WT Pico Kit manual (https://assets.thermofisher.com/TFS-Assets/LSG/manuals/MAN0017893_703262_GeneChipWT_Pico_UG.pdf) as an initial guideline for the number of PCR cycles. The optimal number of PCR cycles should produce 20 to 200 µg of cRNA. Poor-quality or highly degraded samples will require more cycles to get sufficient cRNA yields.
A good practice is to initially run 1-2 samples at a lower number of cycles to determine cRNA yield before deciding to increase the number of cycles for the remaining samples. We recommend keeping the number of cycles consistent within one experiment.
Please note, that running too many cycles will yield a high amount of cRNA, making it difficult to purify and measure concentration. High amounts of cRNA will also cause magnetic beads to become sticky and form loose pellets, making it difficult to separate the beads from the solution.

Find additional tips, troubleshooting help, and resources within our Microarray Analysis Support Center.

We recommend the following assays dependent upon your respective sample size:
- WT Plus Assay for 50-500 ng of total RNA isolated from whole blood, cultured cells, and fresh/fresh frozen tissues.
- WT Pico Assay for 100 pg - 50 ng of total RNA isolated from whole blood, cultured cells, and fresh/fresh frozen or FFPE tissues.

Find additional tips, troubleshooting help, and resources within our Microarray Analysis Support Center.

The following stopping points have been validated for the WT Pico assay.
- After the 14 to 16 hr IVT reaction, the cRNA samples can be stored overnight at -20 degrees C.
- After cRNA purification, the samples can be stored overnight at -20 degrees C. For long-term storage, store samples at -80 degrees C and keep the number of freeze-thaw cycles to 3 or less to ensure cRNA integrity.
- Hydrolyzed ss-cDNA samples can be stored overnight at -20 degrees C.
- Purified ss-cDNA samples can be stored overnight at -20 degrees C. For long-term storage at -20 degrees C, we recommend storing the samples as ss-cDNA and not to proceed to the fragmentation and labeling reaction.
- Fragmented and labeled ss-cDNA samples can be stored overnight at -20 degrees C. For long-term storage, we recommend storing the samples as unfragmented and unlabeled ss-cDNA.
Additional information regarding the assay workflow can be found in the GeneChip WT Pico Reagent Kit User Guide.

Find additional tips, troubleshooting help, and resources within our Microarray Analysis Support Center.

Control Oligo B2 (3 nM) is a pre-labeled DNA spike-in control required for hybridization to the control probes on the array utilized for grid alignment by AGCC. A failure to include Control Oligo B2 in the hybridization cocktail will result in the inability of the software to apply a grid over the scanned image, leading to the unrecoverable loss of sample data from the array. The absence of the Control Oligo B2 also voids the customer's ability to submit an array replacement request.

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The WT Plus kit or the WT Pico kit is recommended for preparing samples for use with the Gene 1.0 ST Array and the Gene 2.0 ST Array.

Find additional tips, troubleshooting help, and resources within our Microarray Analysis Support Center.