Removing Batch Effects from Longitudinal Gene Expression - Quantile Normalization Plus ComBat as Best Approach for Microarray Transcriptome Data.
Technical variation plays an important role in microarray-based gene expression studies, and batch effects explain a large proportion of this noise. It is therefore mandatory to eliminate technical variation while maintaining biological variability. Several strategies have been proposed for the removal of batch effects, although they have not been evaluated in large-scale longitudinal gene expression data. In this study, we aimed at identifying a suitable method for batch effect removal in a large study of microarray-based longitudinal gene expression. Monocytic gene expression was measured in 1092 participants of the Gutenberg Health Study at baseline and 5-year follow up. Replicates of selected samples were measured at both time points to identify technical variability. Deming regression, Passing-Bablok regression, linear mixed models, non-linear models as well as ReplicateRUV and ComBat were applied to eliminate batch effects between replicates. In a second step, quantile normalization prior to batch effect correction was performed for each method. Technical variation between batches was evaluated by principal component analysis. Associations between body mass index and transcriptomes were calculated before and after batch removal. Results from association analyses were compared to evaluate maintenance of biological variability. Quantile normalization, separately performed in each batch, combined with ComBat successfully reduced batch effects and maintained biological variability. ReplicateRUV performed perfectly in the replicate data subset of the study, but failed when applied to all samples. All other methods did not substantially reduce batch effects in the replicate data subset. Quantile normalization plus ComBat appears to be a valuable approach for batch correction in longitudinal gene expression data.
Clinic for General and Interventional Cardiology, University Heart Center Hamburg, Hamburg, 20246, Germany.|Institute of Medical Biometry and Statistics, University Medical Center Schleswig-Holstein, Campus Luebeck, Lübeck, 23562, Germany.|German Center for Cardiovascular Research (DZHK e.V.), partner site Hamburg, Lübeck, Kiel, 20246, Germany.
Christian
C
Müller
Institute of Medical Biometry and Statistics, University Medical Center Schleswig-Holstein, Campus Luebeck, Lübeck, 23562, Germany.|German Center for Cardiovascular Research (DZHK e.V.), partner site Hamburg, Lübeck, Kiel, 20246, Germany.
Arne
A
Schillert
Clinic for General and Interventional Cardiology, University Heart Center Hamburg, Hamburg, 20246, Germany.
Caroline
C
Röthemeier
Institut National de la Santé et de la Recherche Médicale (INSERM), Unité Mixte de Recherche(UMR) enSanté 1166, F-75013, Paris, France.|Institute for Cardiometabolism and Nutrition (ICAN), F-75013 Paris, France.|Sorbonne Universités, Université Pierre et Marie Curie (UPMC Univ Paris 06), UMR_S1166, Team Genomics & Pathophysiology of Cardiovascular Diseases, F-75013, Paris, France.
David-Alexandre
DA
Trégouët
Institut National de la Santé et de la Recherche Médicale (INSERM), Unité Mixte de Recherche(UMR) enSanté 1166, F-75013, Paris, France.|Institute for Cardiometabolism and Nutrition (ICAN), F-75013 Paris, France.|Sorbonne Universités, Université Pierre et Marie Curie (UPMC Univ Paris 06), UMR_S1166, Team Genomics & Pathophysiology of Cardiovascular Diseases, F-75013, Paris, France.
Carole
C
Proust
Institute of Medical Biostatistics, Epidemiology and Informatics (IMBEI) at the University Medical Center of the Johannes Gutenberg University Mainz, Mainz, 55131, Germany.
Harald
H
Binder
Experimental Ophthalmology, Department of Ophthalmology, University Medical Center of the Johannes Gutenberg University, Mainz, 55131, Germany.
Norbert
N
Pfeiffer
Department of Psychosomatic Medicine and Psychotherapy, University Medical Center of the Johannes Gutenberg-University Mainz, Mainz, Germany.
Manfred
M
Beutel
Institute for Clinical Chemistry and Laboratory Medicine, University Medical Center Mainz, Mainz, Germany.
Karl J
KJ
Lackner
Clinic for General and Interventional Cardiology, University Heart Center Hamburg, Hamburg, 20246, Germany.|German Center for Cardiovascular Research (DZHK e.V.), partner site Hamburg, Lübeck, Kiel, 20246, Germany.
Renate B
RB
Schnabel
Institut National de la Santé et de la Recherche Médicale (INSERM), Unité Mixte de Recherche(UMR) enSanté 1166, F-75013, Paris, France.|Institute for Cardiometabolism and Nutrition (ICAN), F-75013 Paris, France.|Sorbonne Universités, Université Pierre et Marie Curie (UPMC Univ Paris 06), UMR_S1166, Team Genomics & Pathophysiology of Cardiovascular Diseases, F-75013, Paris, France.
Laurence
L
Tiret
Preventive Cardiology and Preventive Medicine, Center for Cardiology, University Medical Center Mainz, Mainz, 55131, Germany.|Center for Thrombosis and Hemostasis, University Medical Center Mainz, Mainz, 55131, Germany.|German Center for Cardiovascular Research (DZHK e.V.), partner site Rhine Main, Mainz, 55131, Germany.
Philipp S
PS
Wild
Clinic for General and Interventional Cardiology, University Heart Center Hamburg, Hamburg, 20246, Germany.|German Center for Cardiovascular Research (DZHK e.V.), partner site Hamburg, Lübeck, Kiel, 20246, Germany.
Stefan
S
Blankenberg
Clinic for General and Interventional Cardiology, University Heart Center Hamburg, Hamburg, 20246, Germany.|German Center for Cardiovascular Research (DZHK e.V.), partner site Hamburg, Lübeck, Kiel, 20246, Germany.
Tanja
T
Zeller
Institute of Medical Biometry and Statistics, University Medical Center Schleswig-Holstein, Campus Luebeck, Lübeck, 23562, Germany.|German Center for Cardiovascular Research (DZHK e.V.), partner site Hamburg, Lübeck, Kiel, 20246, Germany.|Center for Clinical Trials, University of Lübeck, Lübeck, 23562, Germany.
Andreas
A
Ziegler
D000328:Adult; D000368:Aged; D005260:Female; D020869:Gene Expression Profiling / Q000379:methods*; D006801:Humans; D008137:Longitudinal Studies; D008297:Male; D008875:Middle Aged; D009000:Monocytes / Q000737:chemistry*; D017711:Nonlinear Dynamics; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D025341:Principal Component Analysis; D011446:Prospective Studies