| QUALITY CONTROL STEPS DESCRIPTION |
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Quality Control Description Type(s):
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dye_swap_quality_control
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Description:
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We examined pairs of arrays in which the tumor sample was labeled with Cy5 and the reference with Cy3 for one array, and the labeling was reversed in the paired array. We analyzed 17 such pairs of arrays for the NCI 2.7K array and 11 such pairs containing clones from the GEM1 8.7K array. Reverse-labeled experiments were performed by using a variety of tumor samples, including prostate tumors from a related study. We computed the average difference in log2 ratio between normalized forward- and reverse-labeled experiments for each clone as an estimate of the residual dye bias for that clone. Of the clones on the NCI 2.7K array, only 7 had an average difference in log2 ratios greater in absolute value than 1 (i.e., 2-fold difference), and for the GEM1 8.7K array, the number was 19. There was no general systematic bias favoring either dye after normalization. For comparing tumor models with each other, the use of an internal reference consistently labeled in all arrays should not result in spurious claims that genes are differentially expressed among models, but may result in missing the detection of differential expression of genes that do not stain adequately with Cy5.
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