title: Rapid Assessment and Visualization of Normality in High-Content and Other Cell-Level Data and Its Impact on the Interpretation of Experimental Results.
abstract: When investigators monitor effects on a population of cells following a perturbation, these events rarely occur in a classical normal (or Gaussian) distribution. A normal distribution is, however, explicitly assumed for events within a single well, in which mean values per well are used as an assay metric and, in general, measures of assay robustness, such as the Z' score and the V factor. Such analysis is not possible for many technologies; however, high-content screening (HCS) measures events of individual cells, which are averaged over the well. These individual cell-level measurements may be analyzed separately. This study quantifies the extent of nonnormality in experimental samples and their effects on determining the EC50 of a test compound and the assay robustness statistics. The results, based on five sets of publicly available data, indicate that the Z' or V-factor score can be improved by as much as 0.44 more than standard calculations, and the EC50 of a dose-response curve can be lowered by as much as fivefold when nonparametric methods are used, but not all data sets show a significant improvement. The effect on analysis depends in part on whether the greatest shift from normality occurs in the upper or lower range of the dose-response curve.
authors.affiliation: Eli Lilly and Company, Lilly Corporate Center, Indianapolis, IN, USA haney_steven_a@lilly.com.
authors.forename: Steven A
authors.initials: SA
authors.lastname: Haney
mesh_terms: D000465:Algorithms; D003627:Data Interpretation, Statistical; D004305:Dose-Response Relationship, Drug; D049452:Green Fluorescent Proteins / Q000737:chemistry; D057166:High-Throughput Screening Assays / Q000379:methods*; D006801:Humans; D061986:MCF-7 Cells; D008962:Models, Theoretical; D016328:NF-kappa B / Q000378:metabolism; D016011:Normal Distribution; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D011381:Programming Languages; D015203:Reproducibility of Results; D018709:Statistics, Nonparametric; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism; D000071557:beta-Arrestins / Q000737:chemistry
chemical_list: D016328:NF-kappa B; D014409:Tumor Necrosis Factor-alpha; D000071557:beta-Arrestins; D049452:Green Fluorescent Proteins; D019869:Phosphatidylinositol 3-Kinases
keywords: cancer and cancer drugs; cell-based assays; high-content screening; image analysis; statistical analyses