A suite of MATLAB-based computational tools for automated analysis of COPAS Biosort data.

Complex Object Parametric Analyzer and Sorter (COPAS) devices are large-object, fluorescence-capable flow cytometers used for high-throughput analysis of live model organisms, including Drosophila melanogaster, Caenorhabditis elegans, and zebrafish. The COPAS is especially useful in C. elegans high-throughput genome-wide RNA interference (RNAi) screens that utilize fluorescent reporters. However, analysis of data from such screens is relatively labor-intensive and time-consuming. Currently, there are no computational tools available to facilitate high-throughput analysis of COPAS data. We used MATLAB to develop algorithms (COPAquant, COPAmulti, and COPAcompare) to analyze different types of COPAS data. COPAquant reads single-sample files, filters and extracts values and value ratios for each file, and then returns a summary of the data. COPAmulti reads 96-well autosampling files generated with the ReFLX adapter, performs sample filtering, graphs features across both wells and plates, performs some common statistical measures for hit identification, and outputs results in graphical formats. COPAcompare performs a correlation analysis between replicate 96-well plates. For many parameters, thresholds may be defined through a simple graphical user interface (GUI), allowing our algorithms to meet a variety of screening applications. In a screen for regulators of stress-inducible GFP expression, COPAquant dramatically accelerated data analysis and allowed us to rapidly move from raw data to hit identification. Because the COPAS file structure is standardized and our MATLAB code is freely available, our algorithms should be extremely useful for analysis of COPAS data from multiple platforms and organisms. The MATLAB code is freely available at our web site (www.med.upenn.edu/lamitinalab/downloads.shtml).

[1]  C. Link,et al.  Direct observation of stress response in Caenorhabditis elegans using a reporter transgene. , 1999, Cell stress & chaperones.

[2]  Aideen Long,et al.  Statistical methods for analysis of high-throughput RNA interference screens , 2009, Nature Methods.

[3]  Marjolein V. Smith,et al.  A Discrete Time Model for the Analysis of Medium-Throughput C. elegans Growth Data , 2009, PloS one.

[4]  J. Vaupel,et al.  A stress-sensitive reporter predicts longevity in isogenic populations of Caenorhabditis elegans , 2005, Nature Genetics.

[5]  O. Zugasti,et al.  Anti-Fungal Innate Immunity in C. elegans Is Enhanced by Evolutionary Diversification of Antimicrobial Peptides , 2008, PLoS pathogens.

[6]  N. Perrimon,et al.  Functional Genomic Analysis of the Wnt-Wingless Signaling Pathway , 2005, Science.

[7]  Marc Ferrer,et al.  Median Absolute Deviation to Improve Hit Selection for Genome-Scale RNAi Screens , 2008, Journal of biomolecular screening.

[8]  T. Lamitina,et al.  Genome-wide RNAi screening identifies protein damage as a regulator of osmoprotective gene expression , 2006, Proceedings of the National Academy of Sciences.

[9]  Peter McCourt,et al.  High-throughput screening of small molecules for bioactivity and target identification in Caenorhabditis elegans , 2006, Nature Protocols.

[10]  Marjolein V. Smith,et al.  Application of a Mathematical Model to Describe the Effects of Chlorpyrifos on Caenorhabditis elegans Development , 2009, PloS one.

[11]  Oliver Hobert,et al.  Automated screening for mutants affecting dopaminergic neuron specification in C. elegans , 2008, Nature Methods.

[12]  H. Cinar,et al.  A method to rank order water soluble compounds according to their toxicity using Caenorhabditis elegans, a Complex Object Parametric Analyzer and Sorter, and axenic liquid media. , 2009, Food and chemical toxicology : an international journal published for the British Industrial Biological Research Association.

[13]  N. Perrimon,et al.  Functional genomics reveals genes involved in protein secretion and Golgi organization , 2006, Nature.

[14]  Marjolein V. Smith,et al.  Medium- and high-throughput screening of neurotoxicants using C. elegans. , 2010, Neurotoxicology and teratology.