Computational protocol: Transcriptomic analysis of monocytes and macrophages derived from CLL patients which display differing abilities to respond to therapeutic antibody immune complexes

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Protocol publication

[…] Total RNA from monocytes and MDCs was extracted with Bioline Isolate II RNA Micro kit in accordance with the prescribed protocol provided with the kit (Bioline). The integrity of the isolated RNA was determined using Agilent RNA 6000 Pico kit (Agilent Technologies) on an Agilent Bioanalyzer. Microarray profiling was performed using Illumina HumanHT-12 v4 Expression BeadChips (Illumina) as per the manufacturer's protocol. Probe intensities have been background corrected using negative control probes and quantile normalized using the neqc function of the limma package.Microarray data have been uploaded to Gene Expression Omnibus under the reference: GSE71409. The original CEL files from GEO accession GSE5099 , were downloaded. These contain gene expression data for freshly isolated monocytes at day 0, macrophages at day 3 and day 7, Interferon Gamma and LPS treated macrophages and IL-4 treated macrophages. These experiments were performed on the Affymetrix Human Genome U133A Array. Arrays from GSE5099 and the current study were preprocessed and normalized with platform specific tools: the Affymetrix arrays were normalized using the RMA algorithm , and the Illumina arrays were quantile normalized after applying the VST algorithm , . Large platform specific batch effects were observed between the two datasets, so an unsupervised empirical Bayes method was used to remove these batch effects after as implemented in the ‘virtualArray’ package . The two datasets were merged on common genes after multiple probes that mapped to the same gene were collapsed by taking the median expression value per sample. Genes that had no expression detected in any sample across both studies (as indicated with a detection p-value < 0.01) were removed from the data. […]

Pipeline specifications

Software tools limma, virtualArray
Databases GEO
Application Gene expression microarray analysis
Organisms Homo sapiens
Diseases Leukemia, Leukemia, Lymphocytic, Chronic, B-Cell