The Affymetrix HGU-133A spike in data set has been used for determining the sensitivity and specificity of various methods for the analysis of microarray data. We show that there are 22 additional probe sets that detect spike in RNAs that should be considered as spike in probe sets. We assign each proposed spiked-in probe set to a concentration group within the Latin Square design, and examine the effects of the additional spiked-in probe sets on assessing the accuracy of analysis methods currently in use. We show that several popular preprocessing methods are more sensitive and specific when the new spike-ins are used to determine false positive and false negative rates.



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