Below are links to various protocols used in the core regarding sample processing.
- Affymetrix IVT Express - This method is used for Affy's 3' based arrays (ie, U219, U133plus2.0, Mouse 430 2.0, etc), starting with 250ng to 400ng total RNA and generating fragmented, biotinylated cRNA.
- Ambion WT Expression - This method is used for Affy's whole transcriptome based arrays (ie, HTAv2, Gene ST arrays, etc), starting with 250ng to 400ng total RNA and generating ds-cDNA.
- Terminal Biotin End-labeling - This method is used in conjunction with the WT expression kits, to fragment and biotinlyate the cDNA prior to hybridiation on Arrays.
Probe labeling protocols:
- Hyb Oven 640- The Affymetrix Hyb Oven is used for maintaning constant temperatures (at 45C, 48C or 50C) during the hybridization step. After probe has been generated and hybridiation mix has been added, samples are hybed for 16 to 24 hours following the manufacturers recommended procedure for each array..
- Fluidics Station 450- The Affymetrix FS 450 is used to wash and stain the hybridized arrays. Each Genechip uses a particular fluidics script provided by the manufacturer.
- AGCC Command Console User Guide- The Affymetrix Scanner 3000 7G is operated with the AGCC software. Once cartridges have been hybed, washed and stained, they are scanned following the manufacturers default settings.
Cartridge based arrays: