We used the Human Genome U133 Plus 2.0 Array (Affymetrix) containing 54,675 probe sets to determine global impacts of A. phagocytophilum infection in NB4 cells at four hours post-infection. The mRNA was isolated using the TRI reagent extraction protocol (Molecular Research Center, Cincinnati) followed by RNeasy cleanup. cRNA production, labeling reactions, hybridization, staining and washing were performed according to the Gene Chip Expression Analysis Technical Manual (Affymetrix, Santa Clara) at the W.M. Keck Facility (Yale University). Image data were quantified by using genechip/suite microarray suite 5.0 (MAS 5.0) and default parameters were used for identification of informative probes. The presence of transcripts was generated according to a detection p-value described in the Statistical Algorithms Reference Guide (Affymetrix, Santa Clara). The one-sided Wilcoxon’s Signed Rank was the statistical method employed to generate detection p-values. Transcripts with a p-value lower than 0.05 were considered present in RNA samples. Global scaling was used to remove differences and facilitate comparison analysis.
