P175Systematic Random Sampling with Virtual Slides: A NewS<strong>of</strong>tware Tool for Tissue ResearchDTreanor 1 ,MDattani 1 ,PQuirke 1 , H Grabsch 11 Pathology and Tumour Biology, <strong>Leeds</strong> Institute <strong>of</strong> Molecular Medicine,University <strong>of</strong> <strong>Leeds</strong>Systematic random sampling is a powerful tool in tissue based research. Itallows accurate quantification <strong>of</strong> tissue elements on pathology slides whileminimising the number <strong>of</strong> samples needed to measure – for example the ratio <strong>of</strong>cancer to stroma, or the distribution <strong>of</strong> blood vessels.Historically optical graticules with conventional light microscopes have beenused for systematic randomised sampling, but this method has significantdrawbacks in terms <strong>of</strong> flexibility and user acceptability. Commercial s<strong>of</strong>tware isavailable which adds grids to captured micrographs, but this requires individualsnapshots <strong>of</strong> tissue to be taken manually. Using virtual slides for this task issimpler, less labour-intensive, and allows recording <strong>of</strong> results directly intocomputer s<strong>of</strong>tware.We describe a system using custom made s<strong>of</strong>tware developed locally whichallows users to select arbitrary areas <strong>of</strong> any virtual slide and add any number <strong>of</strong>sampling points using a systematic grid. <strong>The</strong> system is web based, allowingresearchers to apply a systematic random grid to any sample from their personalcomputer. Once the sampling points are generated they can be viewed locallyusing virtual slide viewing s<strong>of</strong>tware and the scores entered rapidly andefficiently by the user for export and further analysis.P177<strong>The</strong> Digital Pathology Archive: the UMCU initiativePJ Van Diest 11 University Medical Centre Utrecht, Department <strong>of</strong> Pathology, <strong>The</strong>NetherlandsBackground:Our Pathology Department produces yearly about 100.000 slides. After initialdiagnosis, slides <strong>of</strong>ten have to be retrieved. Which takes a lot <strong>of</strong> time, and slidequality deteriorates in time. We have therefore set up infrastructure to create afully digital archive.Methods: Two Aperio scanners processes batches <strong>of</strong> each 120 slides, recognizethe tissue on low resolution, auto focus on a defined number <strong>of</strong> spots, and thenscan the tissue in true colourat x20. Logistics to maximize scanning capacityhave been worked out. File size is about 500 megabytes on average. <strong>The</strong> imagesare stored on a Sun HSM system consisting <strong>of</strong> a storage server, 6 TB fast harddisks and a tape robot <strong>of</strong> 120 TB. Access time for the images varies from a fewseconds for the hard disk to several minutes for the tape storage.Conclusions: It is technically and practically feasible to digitize all slidesproduced by an average size pathology lab, and store them on a SAN. Doingclinicopathological conferences digitally will gradually be implemented whichwill save a lot <strong>of</strong> preparation time and increase flexibility in location <strong>of</strong> theconferences. Further, consultations will be more efficient. Slide images will beintegrated within the Electronic Patient File, and practical slide sessions forteaching purposes will be done digitally using the Slidepath application.P176Tracking with virtual slides: a tool to study diagnostic error inpathologyDTreanor 1 , D Magee 2 , A Bulpitt 2 ,CLim 3 ,PQuirke 11 Pathology and Tumour Biology, <strong>Leeds</strong> Institute <strong>of</strong> Molecular Medicine,University <strong>of</strong> <strong>Leeds</strong>, 2 School <strong>of</strong> Computing, University <strong>of</strong> <strong>Leeds</strong>,3 Department <strong>of</strong> Gastroenterology, Goodhope HospitalBACKGROUND: Diagnostic error in pathology is a significant problem.Studying the reasons for error is difficult because <strong>of</strong> a lack <strong>of</strong> data on thediagnostic process - virtual slides are a useful tool to study error in pathology asthey allow unsupervised study <strong>of</strong> diagnosis.METHODS: Custom s<strong>of</strong>tware was developed to produce visualisations <strong>of</strong> thediagnostic track followed by pathologists as they viewed virtual slides. <strong>The</strong>seshowed the diagnostic path in 3 dimensions, areas studied for >1000ms, andsubject’s comments about the areas viewed. <strong>The</strong> system was used to study 2trainee and 2 expert pathologists diagnosing 60 oesophageal biopsies showingBarrett’s oesophagus. Comparisons <strong>of</strong> the diagnostic tracks between expertsand trainees were studied to classify the reason for errors.RESULTS: 46 cases had an expert consensus diagnosis. <strong>The</strong> two trainees madeerrors in 21 cases (46%) and 15 cases (33%) respectively, <strong>of</strong> which 11 and 9were clinically significant errors. Errors were made across the whole spectrumfrom normal biopsies to intramucosal carcinoma, though there was a tendencyfor serious undercalls to be more common than serious overcalls. Detailedexamination <strong>of</strong> the tracks showed that in all 36 errors there was a error ininterpretation <strong>of</strong> information; in 3 errors there was an additional component <strong>of</strong>failure to identify diagnostic features.CONCLUSIONS: Tracking with virtual slides and the resulting visualisationsare a useful tool in studying diagnosis. <strong>The</strong> tool has the potential for use intraining and assessment in order to improve the quality <strong>of</strong> diagnosis inpathology.P178Non-Side Population Cell Cycle Suppression by Hoechst 33342Occurs in the Absence <strong>of</strong> Significant Cell Death in thePancreatic Cancer Line Capan-2N Guppy 1 , WR Otto 2 , MR Alison 11 Institute for Cell and Molecular Science, Barts and <strong>The</strong> London School <strong>of</strong>Medicine and Dentistry, 2 Histopathology Unit, London Research Institute,Cancer Research UKIsolation <strong>of</strong> a “side population” (SP) via Hoechst (Ho) 33342 efflux assay is apopular tool for adult stem cell identification. High activity <strong>of</strong> ABCtransportersconfers the enhanced dye efflux capacity <strong>of</strong> SP cells and cytotoxicdrug resistance. However, Ho33342 is known to be capable <strong>of</strong> inducingcytotoxic and cytostatic effects, raising the possibility that enhancedclonogenicity and proliferative capacities exhibited by SP cells versus non-SPcells isolated in this way may be an artefact <strong>of</strong> Ho33342 overload in the non-SPpopulation.SP and non-SP cells were isolated from two human pancreaticadenocarcinoma lines (Panc-1 and Capan-2) via standard Ho33342 effluxassay. Viability, colony forming ability (CFA) and proliferation <strong>of</strong> SP, non-SPand unsorted populations exposed to Ho33342 ± reserpine (an inhibitor <strong>of</strong>ABC-transporter activity), reserpine or vehicle alone were examined.Capan-2 SP cells showed elevated CFA versus unsorted Ho-exposedcontrols. However, in unsorted samples, Ho33342 exposure producedsignificant decreases in CFA and population doubling rate, exacerbated by coincubationwith reserpine. Conversely, Panc-1 cells showed no significantdifferences in CFA or proliferation rate between treatments or sub-populations.<strong>The</strong>se differing responses to Ho33342 exposure could not be explained bydifferences in cell cycle position, DNA content or apoptosis levels.Furthermore, the percentage <strong>of</strong> propidium iodide-negative viable cells did notvary between lines or treatments.<strong>The</strong>refore, elevated CFA <strong>of</strong> Capan-2 SP cells may be an artefact <strong>of</strong> dyeinducedsuppression <strong>of</strong> proliferation <strong>of</strong> the non-SP fraction.76 <strong>Summer</strong> <strong>Meeting</strong> (194 th ) 1–4 July <strong>2008</strong> Scientific Programme
AbstractsOralNote: Presenter’s name is shown in bold<strong>Summer</strong> <strong>Meeting</strong> (194 th ) 1–4 July <strong>2008</strong> Scientific Programme77
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