{ "schemaVersion": 4, "meta": { "name": "Example Percent Positive", "shortDescription": "Ported from CellProfiler ExamplePercentPositive pipeline", "description": "Two-channel percent-positive assay (HT29 cells, DNA + phospho-histone H3 mitosis marker). Ported from the CellProfiler ExamplePercentPositive.cppipe example. Channel 0 = OrigBlue/DNA (files *d0.tif), channel 1 = OrigGreen/PH3 (files *d1.tif), merged into one 2-channel TIFF per site via Fiji (images/). Mapping: IdentifyPrimaryObjects -> Gaussian blur (sigma 1.0 = CP threshold smoothing scale 1.3488/0.6744/2) + Threshold computed on the raw image (LI = Minimum Cross-Entropy for nuclei; 3-class Otsu, middle class foreground, lower bound 0.05 for PH3) + connected components + watershed (Shape -> distance-map seeds, Intensity -> intensity seeds with CP's maxima-suppression radius min-diameter/1.5 and the effective sigma 2.83 of CP's truncated 10-px smoothing kernel) + fill object holes (CP: after declumping only) + area filter from the typical diameter (pi*d^2/4) with border objects discarded. RelateObjects + FilterObjects(Children_PH3_Count >= 1) -> class ph3_pos_nuclei (nuclei overlapping a ph3 object). ClassifyObjects(MaxIntensity_OrigGreen > 0.2) -> class ph3_pos_by_intensity. CalculateMath (percent positive) has no pipeline equivalent: compute it after export as count(ph3_pos_nuclei) / count(nuclei) * 100. Result vs CP 4.2.8: nuclei 281 vs 280 (median area 115 vs 118.5 px), PH3 19 vs 20 (median 91 vs 94.5 px), PH3-positive nuclei 20 vs 20, percent positive 7.12 % vs 7.14 %, positive by max intensity 7.47 % vs 7.50 %. Remaining deviations: evanalyzer's distance-map watershed (prominence-based) splits a few large nucleus clumps that CP keeps whole (largest nucleus 354 vs 471 px), and splits one large PH3 object CP keeps whole (largest 121 vs 191 px); CP's 3-class Otsu uses a 128-bin histogram.", "authors": [], "creationTime": "2026-10-21T00:00:00Z", "category": "Assay", "tags": [ "percent-positive", "mitosis", "2-channel", "cellprofiler" ], "appVersion": "0.1.0" }, "classification": { "classes": [ { "id": 0, "color": "#97978f", "name": "Background", "notes": "" }, { "id": 1, "color": "#3366ff", "name": "nuclei", "notes": "Nuclei in CP (DNA channel 0)" }, { "id": 2, "color": "#33ff66", "name": "ph3", "notes": "PH3 in CP (PH3 channel 1)" }, { "id": 3, "color": "#ff3333", "name": "ph3_pos_nuclei", "notes": "PH3PosNuclei in CP: nuclei overlapping at least one ph3 object" }, { "id": 4, "color": "#ff9900", "name": "ph3_pos_by_intensity", "notes": "ClassifyObjects bin PH3Pos in CP: nuclei with max PH3 intensity > 0.2" } ] }, "plate": { "groupingMode": "AUTO", "groupingRegex": "", "plateSize": "AUTO", "wellLayout": "AUTO", "wellImageOrder": [ 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16 ] }, "pipelines": [ { "id": 1, "name": "nuclei", "description": null, "imageSource": { "channel": 0 }, "enabled": true, "steps": [ { "id": 1, "enabled": true, "command": { "type": "GAUSSIAN_BLUR", "kernelSize": 9, "sigma": 1.0 } }, { "id": 2, "enabled": true, "command": { "type": "THRESHOLD", "thresholds": [ { "method": { "type": "LI" }, "minThreshold": 0.0, "maxThreshold": 1.0, "unit": "RELATIVE", "segmentationClass": 1, "valueSource": "RAW_IMAGE" } ] } }, { "id": 3, "enabled": true, "command": { "type": "CONNECTED_COMPONENTS", "minSize": 1 } }, { "id": 4, "enabled": true, "command": { "type": "WATERSHED", "maximumFinderTolerance": 0.5, "smoothingSigma": 0.0, "minObjectSize": 0, "seedSource": "DISTANCE_MAP" } }, { "id": 5, "enabled": true, "command": { "type": "FILL_OBJECT_HOLES" } }, { "id": 6, "enabled": true, "command": { "type": "EXTRACT_OBJECTS", "maxObjectsBeforeFail": 100000, "classMapping": [ { "segmentationClass": 1, "objectClass": { "GLOBAL": 1 } } ] } }, { "id": 7, "enabled": true, "command": { "type": "CLASSIFY_OBJECTS", "originSegmentation": [], "inputClasses": [ { "GLOBAL": 1 } ], "matchHandling": "REMOVE_ALL_CLASSES_IF_NOT_MATCH", "outputClass": "UNSET", "overlappingWith": "UNSET", "minIntersectionArea": 0.0, "sizeUnit": "PIXELS", "minArea": 50.27, "maxArea": 5026.55, "minCircularity": 0.0, "maxCircularity": 1.0, "minSolidity": 0.0, "maxSolidity": 1.0, "minAspectRatio": 0.0, "maxAspectRatio": 2147483600.0, "minEccentricity": 0.0, "maxEccentricity": 1.0, "minFeret": 0.0, "maxFeret": 2147483600.0, "allowEdgeTouching": false, "intensityFilters": [] } } ] }, { "id": 2, "name": "ph3", "description": null, "imageSource": { "channel": 1 }, "enabled": true, "steps": [ { "id": 1, "enabled": true, "command": { "type": "GAUSSIAN_BLUR", "kernelSize": 9, "sigma": 1.0 } }, { "id": 2, "enabled": true, "command": { "type": "THRESHOLD", "thresholds": [ { "method": { "type": "OTSU", "classes": { "type": "THREE", "middleClass": "FOREGROUND" } }, "minThreshold": 0.05, "maxThreshold": 1.0, "unit": "RELATIVE", "segmentationClass": 1, "valueSource": "RAW_IMAGE" } ] } }, { "id": 3, "enabled": true, "command": { "type": "CONNECTED_COMPONENTS", "minSize": 1 } }, { "id": 4, "enabled": true, "command": { "type": "WATERSHED", "maximumFinderTolerance": 3.33, "smoothingSigma": 2.83, "minObjectSize": 0, "seedSource": "INTENSITY" } }, { "id": 5, "enabled": true, "command": { "type": "FILL_OBJECT_HOLES" } }, { "id": 6, "enabled": true, "command": { "type": "EXTRACT_OBJECTS", "maxObjectsBeforeFail": 100000, "classMapping": [ { "segmentationClass": 1, "objectClass": { "GLOBAL": 2 } } ] } }, { "id": 7, "enabled": true, "command": { "type": "CLASSIFY_OBJECTS", "originSegmentation": [], "inputClasses": [ { "GLOBAL": 2 } ], "matchHandling": "REMOVE_ALL_CLASSES_IF_NOT_MATCH", "outputClass": "UNSET", "overlappingWith": "UNSET", "minIntersectionArea": 0.0, "sizeUnit": "PIXELS", "minArea": 19.63, "maxArea": 314.16, "minCircularity": 0.0, "maxCircularity": 1.0, "minSolidity": 0.0, "maxSolidity": 1.0, "minAspectRatio": 0.0, "maxAspectRatio": 2147483600.0, "minEccentricity": 0.0, "maxEccentricity": 1.0, "minFeret": 0.0, "maxFeret": 2147483600.0, "allowEdgeTouching": false, "intensityFilters": [] } } ] }, { "id": 3, "name": "percent positive", "description": null, "imageSource": "SCRATCHPAD", "enabled": true, "steps": [ { "id": 1, "enabled": true, "command": { "type": "CLASSIFY_OBJECTS", "originSegmentation": [], "inputClasses": [ { "GLOBAL": 1 } ], "matchHandling": "ADD_OUTPUT_CLASS_IF_MATCH", "outputClass": { "GLOBAL": 3 }, "overlappingWith": { "GLOBAL": 2 }, "minIntersectionArea": 1.0, "sizeUnit": "PIXELS", "minArea": 0.0, "maxArea": 2147483600.0, "minCircularity": 0.0, "maxCircularity": 1.0, "minSolidity": 0.0, "maxSolidity": 1.0, "minAspectRatio": 0.0, "maxAspectRatio": 2147483600.0, "minEccentricity": 0.0, "maxEccentricity": 1.0, "minFeret": 0.0, "maxFeret": 2147483600.0, "allowEdgeTouching": true, "intensityFilters": [] } }, { "id": 2, "enabled": true, "command": { "type": "CLASSIFY_OBJECTS", "originSegmentation": [], "inputClasses": [ { "GLOBAL": 1 } ], "matchHandling": "ADD_OUTPUT_CLASS_IF_MATCH", "outputClass": { "GLOBAL": 4 }, "overlappingWith": "UNSET", "minIntersectionArea": 0.0, "sizeUnit": "PIXELS", "minArea": 0.0, "maxArea": 2147483600.0, "minCircularity": 0.0, "maxCircularity": 1.0, "minSolidity": 0.0, "maxSolidity": 1.0, "minAspectRatio": 0.0, "maxAspectRatio": 2147483600.0, "minEccentricity": 0.0, "maxEccentricity": 1.0, "minFeret": 0.0, "maxFeret": 2147483600.0, "allowEdgeTouching": true, "intensityFilters": [ { "channel": 1, "metric": "MAX", "comparison": "ABOVE", "threshold": 0.2, "unit": "RELATIVE" } ] } } ] } ], "modules": [] }