OK. After having used beadarray to get very good results from bead-level data, I've recently read all of the work on the multiple artifact removal methods worked out by the Cambridge group (refs at end of email). Indeed, I may have read them a bit too many times. My question is based on figure 5 from the spatial phenomena paper -- the workflow: http://www.biomedcentral.com/content/supplementary/1471- 2105-11-208-s5.pdf -- link to the paper = http://www.biomedcentral.com/1471-2105/11/208 . As of this moment, I have several questions: 1. In which sequence should one peform the steps in the workflow with respect to BASH and HULK? 2. "While BASH removes beads from the analysis, HULK adjusts the values of the beads left after BASHing, and combination of the two tools requires care." -- how should one combine these tools with the material in the spatial phenomena paper? 3. Does one run generateNeighbours before running the supplemental scripts "excludeBrightBeadNeighbours", "nearNonDecodedClusters", and "deviationFromGrid"? 4. How does one set weights for all of those abnormal occurrences? There's no worked example using everything, just individual demonstrations. There's an amazing amount of work (as cited above), and I've already used HULK successfully for several analyses, but the new functions -- they're a bit rough. EBImage's readImage throws up all kinds of flags about the TIFFs -- tags out of order, imagelength missing, etc. They're valid images though, Preview and Photoshop both open them correctly . . . Help is appreciated. Thanks, Anand Refs: Smith ML, Lynch AG. BeadDataPackR: A Tool to Facilitate the Sharing of Raw Data from Illumina BeadArray Studies. Cancer Informatics 2010; 2010:217-27. Smith ML, Dunning MJ, Tavar? S, Lynch AG. Identification and correction of previously unreported spatial phenomena using raw Illumina BeadArray data. BMC Bioinformatics 2010; 11:208. Lynch AG, Smith ML, Dunning MJ, Cairns JM, Barbosa-Morais NL, and Tavare S (2009). beadarray, BASH and HULK - tools to increase the value of Illumina BeadArray experiments. In A. Gusnanto, K.V. Mardia, & C.J. Fallaize (eds), Statistical Tools for Challenges in Bioinformatics, pp.33-37. Leeds, Leeds University Press. Cairns JM, Dunning MJ, Ritchie ME, Russell R, Lynch AG. BASH: A tool for managing BeadArray spatial artefacts Bioinformatics Oct 25 Epub ahead of print [pubmed] Dunning MJ, Barbosa-Morais NL, Lynch AG, Tavar? S, Ritchie ME. Statistical issues in the analysis of Illumina data. BMC Bioinformatics 2008 9:85 Dunning MJ, Thorne NP, Tavar? S. Quality control and low-level statistical analysis of Illumina BeadArrays. REVSTAT - Statistical Journal, Volume 4, Number 1, March 2006, pages 1-30 Anand C. Patel, MD, MS Bioinformatics Assistant Professor of Pediatrics and Medicine Division of Allergy/Pulmonary Medicine Department of Pediatrics Washington University School of Medicine 660 South Euclid Ave, Campus Box 8052 St. Louis, MO 63110 acpatel at wustl.edu acpatel at gmail.com