About 5 ml blood samples were collected, and the PBMCs were isolated using standard density-gradient centrifugation on Ficoll-Paque (MultiSciences (Lianke) Biotech, China). Total RNA was isolated using RNAiso Plus (Takara Bio, Dalian, China) according to
Label
Cy3
Label protocol
Briefly, mRNA was purified from total RNA after removal of rRNA (mRNA-ONLY Eukaryotic mRNA Isolation Kit, Epicentre). Then, each sample was amplified and transcribed into fluorescent cRNA along the entire length of the transcripts without 3' bias utiliz
Hybridization protocol
1 ?g of each labeled cRNA was fragmented by adding 5 ?l 10 × Blocking Agent and 1 ?l of 25 × Fragmentation Buffer, then heated the mixture at 60°C for 30 min, finally 25 ?l 2 × GE Hybridization buffer was added to dilute the labeled cRNA. 50 ?l of hybridi
Scan protocol
The hybridized arrays were washed, fixed and scanned with using the Agilent DNA Microarray Scanner (part number G2505C).
Data processing
Agilent Feature Extraction software (version 11.0.1.1) was used to analyze acquired array images. Quantile normalization and subsequent data processing were performed with using the GeneSpring GX v12.1 software package (Agilent Technologies). After quanti
