Frequently Asked Questions

1 Where is the UMCCC Affymetrix Core Facility located? What is your ship to address
2 What is to be done prior bringing in my samples?
3 What arrangements are necessary in order to drop off samples?
4 What happens if the quality of my RNA is not adequate?
5 What if the concentration, as measured by the Facility, does not match stated requirements?
6 What are my starting material options, how much RNA?
7 When can I expect my results back?
8 What happens to any leftover starting material, cRNA and/or probe?
9 What sort of data does the facility provide me with?
10 Can I obtain help with the analysis of my data?
11 What happens if I get a defective chip?
12 How do I know when I have a defective chip?
13 How often does the Core offer training seminars?
14 If I make my own probes, what quality checks should I perform?
15 My cRNA yields are consistently low, though my total RNA quality appears to be very good. What is the problem?
16 Can you probe with less than 15ug of cRNA?
17 Can I buy chips from the Core? Can I buy labeling kits?
18 I have leftover chips; can I sell them back to you or Affymetrix?
19 I plan to synthesize my own probes, where can I get protocols/advise?

1- The UMCCC Affymetrix Core Facility is located at the NCRC, Building 14, Room 148. Our ship to address is: 2800 Plymouth Rd, NCRC Bldg 14, Room 148, Ann Arbor, MI 48109-2800
2- You must register and submit your sample information using the appropriate form: Guidelines for sample submission.
3- Call (7-5623 or 4-1461) or Email in order to arrange for a drop off date and time. Please do not just show up and leave samples with people who are not members of the Core Lab. The main campus drop off site is at 2568 MSRB II, 4-1461
4- Samples should be checked before submitting to the core. Please note that, if you ask us to process a sample we deem to be of questionable quality, we are NOT responsible for poor yields in probe synthesis. A second probe synthesis will NOT be done free of charge.
5- We are aware that slight variations will occur when different people are measuring concentrations. Unless the differences between the concentrations you submit and the ones we obtain are significant, we will proceed with the processing of the sample. If there is a significant discrepancy, you will be contacted via email. If we are obligated to manipulate the sample in order to attain a desirable working concentration (speed-vac/precipitation) a $30 surcharge per sample will apply.
6- We typically get very good results with 500ng of total RNA as starting material using the standard Affy processing protocol. We are now also processing smaller amounts, 10ng, using the Ovation Biotin Labeling system or WT-Pico v2 system from NuGen, Inc. All methods generate enough labeled probe for hybridization to Affymetrix GeneChips.
7- Samples are handled on a first come first served basis. Usually, if you bring in total RNA, your samples will be ready in 3-4 weeks. If you bring in fragmented cRNA labeled probe, 1 week.
8- The Core will store your starting material, cRNA and probe at –20 C for up to a month. We are NOT responsible for materials left with us for more than a month. MAKE SURE TO COLLECT YOUR RNA, CRNA AND PROBES WHEN YOU PICK UP YOUR DATA!!! PROBES CAN BE REUSED!!! CDNA CAN BE USED TO MAKE MORE PROBES!!!
9- The Core will extract data from individual chips for you and provide a report for you customized to your experimental questions. Please arrange a time to meet with Craig Johnson, the core biostatistian (7-5623) to discuss your experiment. Upon request, you will get DAT (image), CHP (data in Affymetrix software format), CEL (averaged intensity), TXT (data in text format, tab limited, same information as CHP) and EXP (general experiment info) files from us. We do not store data indefinately, thus you are solely responsible for obtaining/storing data files.
10- The Core provides users with basic data analysis. Craig Johnson, UMCCC Microarray Core Statistician, will preform a preliminary data analysis. More rigorous techniques are also available.
11- Defective chips are replaced by Affymetrix free of charge. Replacement of a chip can take up to a month. Contact Affymetrix to arrange replacement. Typically, Affymetrix will request a copy of the DAT and EXP files for the defective chip. Occasionally, they have requested the chip itself.
12- We routinely check for poor chip performance and will be able to tell you when a redo will be required.
13- Affymetrix may provide us with training seminars periodically. Affymetrix staff holds the training sessions at the Cancer Center. This sessions will typically address issues such as data analysis using Affymetrix software, how to use the software, etc. Anouncements of training will be posted on the website and communicated to current users.
14- Check the quality and purity of your starting material. Your A260/A280 ratio should be >1.7 if you dilute your sample in 10mM Tris pH8 and >1.4 if diluting in H2O. A quality estimate is done by running a gel and getting the 28S to 18S rRNA band ratio. It should be about 2, at the very least 1.6. Merely eyeballing a gel to see if your 28S and 18S bands look OK is not adequate. Measure your cRNA yield, you should routinely obtain >50ug cRNA if starting from 10ug total RNA (this is after adjusting your yield by subtracting starting RNA material). Run your cRNA in a gel to make sure that you have a smear distributed from ~1kb to ~6kb. After fragmentation, run another gel to make sure that your cRNA has been properly fragmented, sizes should range from 100 to 500 bp.  
15- Check the kit troubleshooting guide. Likely when you are setting up your IVT reaction you are not letting the components warm up to room temperature. You should let ALL the components of the reaction (including cDNA and with the exception of the enzyme) warm up to room temperature. The reaction buffer contains spermidine, which can precipitate cDNA in cold solutions.
16- The Core has conducted hybridizations with as low as 12.5 ug of cRNA with some systematic deviations as compared to 15ug of cRNA. We recommend 15ug; it is your decision. If you decide to use lower or higher amounts you should keep that consistent among all of your experiments if you plan on comparing chips from different experiments.
17- You can purchase Affy GeneChips from the Core, however, we do not sell any labeling kits or components.
18- No, that is not possible. We are sensitive to the fact that these are expensive/perishable items and research money is tight. The best thing that we can offer at this point is to use the Medical Schools email list server to allow people to either exchange chips or pay via interdepartmental vouchers. Alternatively, researchers not needing all the chips available in a box can team up with another lab so that all chips get used upon purchase.
19- You may request protocols directly from Affymetrix.

Review the GeneChip overview page for additional information.