Venous blood was collected by simple venipuncture under aseptic conditions. All samples were processed within two hours of collection to minimize gene expression variations associated with longer sample incubation times. PBMCs were separated by Ficoll density gradient, and were immediately lysed in Trizol reagent (Invitrogen, Carlsbad, California). Additional purification was performed on RNeasy columns (Qiagen, Valencia, CA 913555, cat. no. 74104). The quality of total RNA samples was assessed using an Agilent 2100 Bioanalyzer (Agilent Technologies, Palo Alto, CA).
Label
biotin
Label protocol
Total RNA was labled with the Ambion TotalPrep RNA Amplification Kit according to manufacturers suggested protocols.
Hybridization protocol
Standard Illumina hybridization protocol
Scan protocol
Standard Illumina scanning protocol
Description
Scleroderma only
Data processing
A single intensity (expression) value for each Illumina probe on the array was obtained using Illumina BeadStudio software with standard settings and no background correction. The expression values for all the probes for each sample were scaled to have median 256 (28) and then log (base 2) transformed before performing statistical analysis.
