Expression data from draining lymph node of mice immunized with AS01 or its components

Immunization with adjuvanted antigen results in coordinate changes in gene expression. Combination adjuvants exploit the interplay between different immunostimulants to promote immune response to vaccination. The Adjuvant System 01 (AS01) is a liposome-based combination adjuvant which contains two immunostimulants, MPL and QS-21 Here, we assess the contribution of the components of AS01 on its immunogenicity by comparing the transcriptional response induced in the draining lymph node by AS01, MPL and QS-21 at 2,4 or 6h after intramuscolar (i.m.) immunization C57BL/6 mice (6-8 weeks old) were immunized in both hind limbs, in the gastrocnemius (gcm) muscles and in a volume of 50 µl/muscle with AS01b (100mg/ml QS-21 + 100mg/ml MPL) or QS-21 (100mg/ml) or MPL (100mg/ml), all formulated in liposomes. Ileac lymph nodes were identified as draining in setup experiments. Total RNA was isolated by homogenizing pooled dLN in Tripure reagent (1 ml/100 mg tissue; Roche Applied Science) and then extracted with chloroform followed by RNeasy™ Minikit (Qiagen) according to the manufacturer’s protocol. A DNAse treatment was applied on the RNeasy column to avoid genomic DNA contamination. RNA was concentrated by ethanol precipitation, and quantified by RiboGreen™ (Life Technologies). 1 g of each RNA sample was used for target preparation,using a one-cycle cDNA synthesis kit, and hybridized to GeneChip_ Whole Mouse Genome 430 2.0 arrays (Affymetrix). Data acquisition was performed using GeneChip Operating Software (Affimetrix). For each condition, 6 mice were treated. dLN from 2 mice were pooled for each hybridization. 3 replicate chips were analysed per condition.