FAQ
Affymetrix Microarrays:
- What is the Affymetrix GeneChip array?
- What is the price of Affymetrix arrays?
- What Affymetrix microarrays are available? Where can I get a genelist for available arrays?
- How many samples do I need to prepare for an Affymetrix microarray?
After you get your results:
- What options do I have for data analysis after I get my results?
- What are the result files for an Affymetrix array? Which one should I use for which analysis program?
Affymetrix Microarrays:
What is the Affymetrix GeneChip array?
Answer: The Affymetrix GeneChip system provides an approach to comparatively analyze genome wide patterns of gene expression using a technology that incorporates miniaturized, high density arrays of 25mer oligonucleotide probes. The probe arrays are manufactured by Affymetrix's proprietary, light directed chemical synthesis process, which generates high density arrays of oligonucleotides that possess a predefined position on the array. These arrays are used to monitor gene expression for thousands of transcripts. A transcript is represented as a probe set. A probe set is made up of probe pairs comprised of a perfect match (PM) and a mismatch (MM) probe cells. This probe pairing strategy identifies and minimizes the effects of non specific hybridization and background signal. The intensities of each probe pair are used to determine the expression measurement. This measurement is calculated for each probe set and is described in the form of qualitative and quantitative values using the Affymetrix GeneChip Command Console software, v1.1.
Briefly, target preparation involves starting with 2 micrograms of total RNA from tissue or cells. An invitro transcription reaction is then performed to produce a biotin-labeled cRNA from the cDNA. The cRNA is fragmented before hybridization and a hybridization cocktail is prepared that contains the fragmented cRNA, probe array controls, BSA and herring sperm DNA. The cRNA is hybridized to the oligonucleotide probes on the array for 16 hours at 45C. Immediately following hybridization, the hybridized probe arrays undergo an automated washing and staining protocol on the fluidic station and are then scanned on the GeneChip Scanner 3000 7G where patterns of hybridization are detected. The hybridization data are colleced as light emitted from the fluorescent reporter groups already incorporated into the target, which is now bound to the probe array. Probes that perfectly match the target generally produce stronger signals than those that have mismatches. The scanner acquires an image of each of the probe cells and the computer workstation automatically overlays two scanned images and averages the intensities of each probe cell for the greatest array sensitivity. Data generated from the scan is then analyzed using the Affymetrix GeneChip Command Console software, v1.1.
What is the price of Affymetrix arrays?
Answer: Duke investigators must purchase arrays directly from Affymetrix and bring them to the facility when submitting samples. Based on the volume of use, Duke investigators receive a considerable discount on the price of Affymetrix GeneChip arrays. For information on array pricing, please contact us. To determine the costs for probe synthesis, hybridization, and analysis of Affymetrix arrays, please click here.
What Affymetrix microarrays are available? Where can I get a genelist for available arrays?
Answer: Go to www.affymetrix.com to view the various genechips available. Go to "Additional Support" under each chip to download genelists.
How many samples do I need to prepare for an Affymetrix microarray?
Answer: Only the sample of interest is necessary (a reference sample is not used). The sample should be >2μg of total RNA in 8μl RNase-free water; 1-3μg of mRNA in 10μl RNase-free water; 20μg of fragmented cRNA in 40μl. For total RNA we also perform an RNA quality check, so please bring an extra 3μl aliquot of your sample in a seperate tube.
After you get your results:
What options do I have for data analysis after I receive the results?
Answer: If you choose to do your own analysis, we can help you with any supported software listed on our Data Analysis Page.
We can also perform data analysis for you. Click here for more information.
What are the result files for an Affymetrix array? Which one should I use for which analysis program?
Answer: For each array experiment that is performed using the Affymetrix GeneChip arrays, six files are generated (*.DAT, *.CEL, *.CHP, *.ARR, *.RPT, *.txt). Each file will be named identically except for the extension. The naming convention is as follows:
Project ID number_Submission ID nubmer_Genome ID number_sample name_chip type, i.e. 1234_0001_12123_CX24R_HU133+2.*
In the case of pairwise comparisons in the Affymetrix Microarray Suite v5.0, the comparison files will follow the following naming convention:
Sample_base_Sample_exp.txt
The types of extensions are as follows:
| Extension | Description | Comments |
| *.DAT | Scanned image of the GeneChip array | Can only be opened in Affymetrix GeneChip Command Console |
| *.CEL | Cell intensity file that calculates the average intensities for each cell and assigns it to an x,y coordinate position | Can be opened in Excel to manipulate |
| *.CHP | Contains analysis output | Can only be opened in Affymetrix GeneChip Expression Console |
| *.ARR | Contains experimental information | Can only be opened in Affymetrix GeneChip Expression Console |
| *.RPT | Contains quality control information about the chip | Can be opened in Excel to manipulate |
| *.txt | Contains analysis output | Can be opened in Excel to manipulate |
All data analyses will be given as *.txt files.