Bossers et al. BMC Genomics 2010, 11:112 http://www.biomedcentral.com/1471-2164/11/112 separate arrays, and against different samples. For exam- ple, in dataset C1, the directly measured ratios between samples T1 and T2 on array 1, were compared with the reconstructed ratios between T1 on array 4, and T2 on array 2 (Additional file 1). To eliminate possible con- founding effects of noise introduced by genes expressed at very low levels, only genes with an average log-trans- formed intensity levels greater than 7 were used. To compare the overlap between gene rankings based on the ratio and intensity models, genes were ordered by p- value and assigned to bins containing 1,000 genes. The fraction of overlap then was defined as the amount of genes ranked in the same bin by both models, divided by the size of the bin. Both for the cell line datasets C1 and C2, and the human brain datasets B1 and B2, reproducibility between the replicated datasets was determined as fol- lows. First, the correlation between sets of p-values was calculated using Spearman’s rho. Second, to assess the proportion of genes with similar ranks between repli- cates, genes were ordered by p-value. For bins of increasing size (10 to 1,000 genes, by increments of 10 genes), the proportion of overlap was defined as the fraction of genes, occurring in both sets. Third, Bayesian information criterion (BIC) model selection was used to score the ratio- and intensity-based linear models for each array feature. Information criterion methods, which aim to determine which set of model parameters the data support best, penalize models with more unknown parameters in order to select a model with a lower gen- eralization error and hence more reproducible results [15]. The preferred model was defined by the model with the lowest BIC value. BIC calculations were per- formed using the nlme package in R. Additional file 1: Hybdridization setup cell line experiment. DN = P53DN mutant, SH= shRNA, MI = MIR372, E6 = HPV16 E6. The array numbers are given in the top left corner of each array. Click here for file [ http://www.biomedcentral.com/content/supplementary/1471-2164-11- 112-S1.PNG ] Additional file 2: Correlation real and in silico reconstructed ratios. Correlation between ratios, as measured on the array, and the in silico reconstructed ratios of the same sample pair, as measured on different arrays and against different co-hybridized samples. Only genes with an average log2-transformed expression > 7 were included. Click here for file [ http://www.biomedcentral.com/content/supplementary/1471-2164-11- 112-S2.XLS ] Additional file 3: Correlation real versus virtual hybridizations. Correlation between M-values derived from real hybridizations versus M- values derived from virtual hybridizations for the MAQC dataset. Main title: hybridization number. X-axis: directly measured ratio between sample A and B. Y-axis: mean M-value between 9 virtual comparisons sample A versus sample B, where sample A and B where measured on different arrays. Correlation: Pearson’s correlation coefficient. Click here for file [ http://www.biomedcentral.com/content/supplementary/1471-2164-11- 112-S3.ZIP ] Page 12 of 13 Additional file 4: Hybridization setup human brain experiment. The column dataset denotes in which replicate dataset (B1 or B2) the intensity measurement was used. Click here for file [ http://www.biomedcentral.com/content/supplementary/1471-2164-11- 112-S4.XLS ] Additional file 5: Reproducibility of between-group treatment effects based on ratio and intensity models. Reproducibility of ANOVA-derived treatment effects between group 2 and group 4, and group 1 and group 5, in replicate brain datasets B1 and B2. Reproducibility is enhanced when using intensity-based models instead of ratio-based models. Click here for file [ http://www.biomedcentral.com/content/supplementary/1471-2164-11- 112-S5.PDF ] Additional file 6: Hybdridization setup brain experiment. BR = Braak stage. Click here for file [ http://www.biomedcentral.com/content/supplementary/1471-2164-11- 112-S6.PDF ] Additional file 7: Human keratinocyte cell line dataset. Normalized expression values and raw hybridization data for all hybridizations have been deposited into the Gene Expression Omnibus (GEO) database. Reviewer link GEO dataset: http://www.ncbi.nlm.nih.gov/geo/query/acc. cgi?token=djkxjoomeyecqja&acc=GSE12553. Click here for file [ http://www.biomedcentral.com/content/supplementary/1471-2164-11- 112-S7.ZIP ] Acknowledgements Funding for the human keratinocyte cell line experiment was provided by the EU Research Sixth Framework programme, project DISMAL, project number LSHC-CT-2005-018991. The human brain experiment received funding from the Royal Dutch Academy of Sciences, Innovation Fund and Solvay Pharmaceuticals. Author details 1Neuroregeneration Laboratory, Netherlands Institute for Neuroscience, Meibergdreef 47, 1105 BA Amsterdam, the Netherlands. 2Microarray Core Facility, Department of Pathology, VU University Medical Center, PO Box 7075, 1007 MB Amsterdam, the Netherlands. 3Section of Tumour Biology, Department of Otolaryngology/Head-Neck Surgery, VU University Medical Center, De Boelelaan 1117, 1081 HV Amsterdam, the Netherlands. 4Department of Mathematics, Vrije Universiteit, De Boelelaan 1081a, 1081 HV Amsterdam, the Netherlands. 5Department of Biostatistics, VU University Medical Center, PO Box 7075, 1007 MB Amsterdam, the Netherlands. Authors’ contributions KB generated the human brain dataset, conceived of the study, performed statistical analyses, and drafted the manuscript. RHB and SJS generated the human keratinocyte cell line dataset. BY and JV participated in the design of the study and helped to draft the manuscript. MAW conceived of the study, performed statistical analyses and helped to draft the manuscript. All authors read and approved the final manuscript. Competing interests The authors declare that they have no competing interests. Received: 8 October 2008 Accepted: 17 February 2010 Published: 17 February 2010 References 1. Blalock EM, Geddes JW, Chen KC, Porter NM, Markesbery WR, Landfield PW: Incipient Alzheimer’s disease: microarray correlation analyses reveal major transcriptional and tumor suppressor responses. Proc Natl Acad Sci USA 2004, 101:2173-2178.