gender: Female tissue: RAT Peripheral blood mononuclear cells strain: Wistar rat subject group: RA rats after 1 hour ANE therapy
Treatment protocol
Peripheral blood mononuclear cells (PBMC) were freshly isolated on Ficoll gradient (Histopaque®-1077, Sigma Aldrich, USA) according to the manufacture procedures. Separated cell were maintained in RNA Later (Ambion, USA) at -80ºC until the end of the study and then processed in a whole batch for the following extraction.
Growth protocol
Blood sampling was carried out before any therapy in RA (RA_T0) and NORA (NORA_T0) groups whereas in the other groups (AP/MEC, MTX,ANE, PLA and NORA) at 1hour and at 34 days after therapy. Animals were induced in deep general anesthesia and after total blood drawing (5ml), sacrificed by neck dislocation.
Extracted molecule
total RNA
Extraction protocol
Total RNA was extracted and purified using mirVanaTM mRNA and miRNA Isolation Kit (Cat#AM1560, Ambion, Austin, TX, US) checked for the RIN number to inspect RNA integrity by an Agilent Bioanalyzer 2100 (Agilent technologies, Santa Clara, CA, US).
Label
biotin
Label protocol
Total RNA were amplified, labeled and purified by using GeneChip 3’IVT Express Kit (Cat#901229, Affymetrix, Santa Clara, CA, US) followed the manufacturer’s instructions to obtain biotin labeled cRNA.
Hybridization protocol
Array hybridization and wash was performed using GeneChip® Hybridization, Wash and Stain Kit (Cat#900720, Affymetrix, Santa Clara, CA, US)in Hybridization Oven 645 (Cat#00-0331-220V, Affymetrix, Santa Clara, CA, US) and Fluidics Station 450 (Cat#00-0079, Affymetrix, Santa Clara, CA, US) followed the manufacturer’s instructions.
Scan protocol
Slides were scanned by GeneChip® Scanner 3000 (Cat#00-00212, Affymetrix, Santa Clara, CA, US) and Command Console Software 3.1 (Affymetrix, Santa Clara, CA, US) with default settings. Raw data were normalized by MAS 5.0 algorithm, Gene Spring Software 11.0 (Agilent technologies, Santa Clara, CA, US).
Description
Peripheral blood mononuclear cells from ANE after 1 hour therapy, replicate 2 Gene expression data from RA rats after 1 hour ANE therapy
Data processing
Raw data Affymetrix .CEL files, were pre-processed using robust multi-array average (rma) in the affy R package.
