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PHLIP quantitative biofilm imaging

PHLIP quantitative biofilm imaging. Why another imaging software?. Group of Prof. J. S. Almeida, ITQB, Lisbon. modern confocal microscopes record fluorescent signals in parallel for multiple channels automated image analysis required to process large datasets

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PHLIP quantitative biofilm imaging

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  1. PHLIPquantitative biofilm imaging

  2. Why another imaging software? Group of Prof. J. S. Almeida, ITQB, Lisbon • modern confocal microscopes record fluorescent signals in parallel for multiple channels • automated image analysis required to process large datasets • a software package which is extensible and flexible

  3. How PHLIP started… Group of Prof. J. S. Almeida, ITQB, Lisbon • previous work of Joao Xavier in image analysis - Xavier, J. B et al. 2001. Objective threshold selection procedure (OTS) for segmentation of scanning laser confocal microscope images. J Microbiol Methods 47:169-80. - Xavier, J. B et al . 2003. Automated biofilm morphology quantification from confocal laser scanning microscopy imaging. Water Sci Technol 47:31-7. • 2003: EU-Project PHOBIA multi-channel CLSM software: PHOBIA Laser scanning Image Processor • Joao unifies his image analysis work in a MATLAB package; version 0.1 of PHLIP • 2004: start in PHOBIA and continue developing PHLIP, current release is version 0.4.1

  4. What is PHLIP? Group of Prof. J. S. Almeida, ITQB, Lisbon • MATLAB package, no additional toolboxes needed • modular architecture, can be easily modified or extended by new imaging function • features a new XML data structure PHLIP-ML • detailed information on: http://www.phlip.org • freely available under an open source license (http://sourceforge.net/projects/phlip)

  5. Program flow of PHLIP Group of Prof. J. S. Almeida, ITQB, Lisbon • Matlab package, no additional toolboxes needed • freely available under an open source license: http://sourceforge.net/projects/phlip • supports different microscope Leica formats • features a new XML data structure, PHLIP-ML

  6. 1. Data Input Group of Prof. J. S. Almeida, ITQB, Lisbon • support for different Leica formats • easily extendable with other formats • PHLIP-ML, a new vendor neutral CLSM format

  7. 2. Image preprocessing Group of Prof. J. S. Almeida, ITQB, Lisbon Several function to preprocess the image data:

  8. thresholded threshold & original 3. Thresholding and GUI Group of Prof. J. S. Almeida, ITQB, Lisbon Segmentation of the images can be done by: - automatic threshold determination - manually with help of GUI

  9. 4. Morphological description Group of Prof. J. S. Almeida, ITQB, Lisbon - Single channel: most popular parameters (Yang et al., 2000) - Two channel: comparative analysis (Xavier et al., 2003) - All channel: overall description of fluorescent channels

  10. 5. Data output Group of Prof. J. S. Almeida, ITQB, Lisbon • Save results in HTML formatted file… • …or export in PHLIP-ML data format:

  11. Application of PHLIP Group of Prof. J. S. Almeida, ITQB, Lisbon PHLIP was presented at the annual IWA Biofilm Conference, October 2004. • publications: • Mueller et al., 2004, Proceedings IWA Biofilm symposium. Las Vegas, Montana. • GrayMerod et al., 2004, Proceedings IWA Biofilm symposium. Las Vegas, Montana. • Mueller et al., submitted. • some research groups working with PHLIP: • Prof. S Wuertz, UC Davis, USA • Dr. JFC de Brouwer, Yerseke, NL • Dr. T Neu, Magdeburg, D

  12. Bacteria Extracel. matrix Algae Application of PHLIP Group of Prof. J. S. Almeida, ITQB, Lisbon Example application of de Brouwer (unpublished): -> development of phototrophic biofilm at different light intensities -> biovolume analysis of 3 channels: bacteria / extracel. matrix / algae's

  13. Outlook Group of Prof. J. S. Almeida, ITQB, Lisbon • new morphological parameters as for example Fractal Dimension in 3D • support for CLSM data from cryosections • function for X / Y - rotation of shifted image stacks

  14. Acknowledgements Group of Prof. J. S. Almeida, ITQB, Lisbon Thanks to… • Jonas Almeida • Joao Xavier • Jody de Brouwer • All members of the Biomath Group in Lisbon

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