title: Assessment of the relationship between pre-chip and post-chip quality measures for Affymetrix GeneChip expression data.
abstract: BACKGROUND
  Gene expression microarray experiments are expensive to conduct and guidelines for acceptable quality control at intermediate steps before and after the samples are hybridised to chips are vague. We conducted an experiment hybridising RNA from human brain to 117 U133A Affymetrix GeneChips and used these data to explore the relationship between 4 pre-chip variables and 22 post-chip outcomes and quality control measures.
  
  
  RESULTS
  We found that the pre-chip variables were significantly correlated with each other but that this correlation was strongest between measures of RNA quality and cRNA yield. Post-mortem interval was negatively correlated with these variables. Four principal components, reflecting array outliers, array adjustment, hybridisation noise and RNA integrity, explain about 75% of the total post-chip measure variability. Two significant canonical correlations existed between the pre-chip and post-chip variables, derived from MAS 5.0, dChip and the Bioconductor packages affy and affyPLM. The strongest (CANCOR 0.838, p < 0.0001) correlated RNA integrity and yield with post chip quality control (QC) measures indexing 3'/5' RNA ratios, bias or scaling of the chip and scaling of the variability of the signal across the chip. Post-mortem interval was relatively unimportant. We also found that the RNA integrity number (RIN) could be moderately well predicted by post-chip measures B_ACTIN35, GAPDH35 and SF.
  
  
  CONCLUSIONS
  We have found that the post-chip variables having the strongest association with quantities measurable before hybridisation are those reflecting RNA integrity. Other aspects of quality, such as noise measures (reflecting the execution of the assay) or measures reflecting data quality (outlier status and array adjustment variables) are not well predicted by the variables we were able to determine ahead of time. There could be other variables measurable pre-hybridisation which may be better associated with expression data quality measures. Uncovering such connections could create savings on costly microarray experiments by eliminating poor samples before hybridisation.
authors.affiliation: Depts. of Psychological Medicine and Medical Genetics, School of Medicine, Cardiff University, Cardiff, UK. jonesl1@cf.ac.uk
authors.forename: Lesley
authors.initials: L
authors.lastname: Jones
authors.forename: Darlene R
authors.initials: DR
authors.lastname: Goldstein
authors.forename: Gareth
authors.initials: G
authors.lastname: Hughes
authors.forename: Andrew D
authors.initials: AD
authors.lastname: Strand
authors.forename: Francois
authors.initials: F
authors.lastname: Collin
authors.forename: Stephen B
authors.initials: SB
authors.lastname: Dunnett
authors.forename: Charles
authors.initials: C
authors.lastname: Kooperberg
authors.forename: Aaron
authors.initials: A
authors.lastname: Aragaki
authors.forename: James M
authors.initials: JM
authors.lastname: Olson
authors.forename: Sarah J
authors.initials: SJ
authors.lastname: Augood
authors.forename: Richard L M
authors.initials: RL
authors.lastname: Faull
authors.forename: Ruth
authors.initials: R
authors.lastname: Luthi-Carter
authors.forename: Valentina
authors.initials: V
authors.lastname: Moskvina
authors.forename: Angela K
authors.initials: AK
authors.lastname: Hodges
mesh_terms: D016477:Artifacts; D003198:Computer Simulation; D003627:Data Interpretation, Statistical; D004867:Equipment Design; D019544:Equipment Failure Analysis; D020869:Gene Expression Profiling / Q000295:instrumentation*; D008957:Models, Genetic; D015233:Models, Statistical; D020411:Oligonucleotide Array Sequence Analysis / Q000295:instrumentation*; D011785:Quality Assurance, Health Care / Q000379:methods*; D011786:Quality Control; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity