P-MTAB-28526 - P-MTAB-28526
Hybridization of labeled cRNA to the BeadChip, and washing and scanning were performed according to the Illumina GeneExpression Direct Hybridization Manual manual. Essentially, the amplified, biotin-labeled human cRNA samples were resuspended in a solution of Hyb E1 buffer (Illumina) and 25% (v/v) formamide at a final concentration of 25 ng/µL. 1.5 µg of each cRNA were hybridized. Hybridization was allowed to proceed at 58 C for 16 hours, after which, the bead array matrix was washed for 10 minutes with 1X High temperature buffer (Illumina), followed by a subsequent 10 minute wash in Wash E1BC buffer. The arrays were then washed with 100% ethanol for 10 min to strip off any remaining adhesive on the chip. A 2 minute E1BC wash was performed to remove residual ethanol. The array signal was developed via 10 minute incubation in Block E1 buffer, containing Streptavidin-Cy3 at a final concentration of 1µg/mL. The BeadChip was washed a final time in Wash E1BC buffer for five minutes and subsequently dried via centrifugation for 4 minutes at a setting of 275 rcf at room temperature.