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In these situations, a direct-comparison design helps to maximize signal-to-noise ratios in the ratios between the two groups through minimization of the array effect and the possibility for more replicates with the same number of arrays.
Model 1 represents the ratio-based analysis: (1) Here, μ captures the average gene intensity, τ i is the treatment specific effect, η j is the cell line (10 or 19) effect, α k (j ) is the array effect, and ε ijk is the error component.
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We first verified that there are intensity-dependent array effects and effect due to the two batches.
The probe intensities for the third simulation scenario included biological effects, array effects and intensity-dependent dye effects as described above.
Meanwhile the standardized estimates of differential expression (y i ) are adjusted for all other sources of variation fitted in the within-study model (i.e. dye and array effects), and standardized by the standard error of the estimate.
To apply SNM, all terms in the model were defined exactly as in the first scenario, except we now defined f 1(BX + AZ) to be a 100 000 × 12 matrix that represents the intensity-dependent array effects and f 2(BX + AZ) to be a 100 000 × 12 matrix that represents the intensity-dependent dye effects.
Affymetrix raw data files (cell intensity [CEL] files) were first analyzed with robust multi-array Average (RMA) normalization as implemented in the Affymetrix Expression Console Software (version 1.1) to remove between-array effects and to standardize the low-level data [ 62].
After applying multiple background, intra- and inter-array effect corrections and signal extraction and probe summarisation techniques, we demonstrate the factors affecting the concordance of both magnitude and statistical significance of the transcriptomic effects between platforms.
Again, two ANOVA models were used: the ratio model which included the array effect α k (j ), and the intensity model without array effect.
We therefore, for the cell line dataset, compared the ranking of genes by p-values generated with two different ANOVA models: one including the array effect (the ratio analysis), and one without the array effect (the intensity analysis).
When intensities instead of ratios were taken into the statistical analysis, no differential genes were detected at a FDR of 10% unless dye and array effect were included in the model.
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