Saved Pipeline, in file Dobbelaere2.txt, Saved on 30-Jul-2008 Pixel Size: 1 Pipeline: LoadImages RescaleIntensity Resize RescaleIntensity Resize RescaleIntensity Resize IdentifyPrimAutomatic IdentifyPrimAutomatic IdentifySecondary IdentifyPrimAutomatic MeasureObjectAreaShape FilterByObjectMeasurement MeasureObjectIntensity FilterByObjectMeasurement Relate Relate Relate SpeedUpCellProfiler Module #1: LoadImages revision - 2 How do you want to load these files? Text-Exact match Type the text that one type of image has in common (for TEXT options), or their position in each group (for ORDER option): s1_w1.tif What do you want to call these images within CellProfiler? Blue Type the text that one type of image has in common (for TEXT options), or their position in each group (for ORDER option): s1_w2.tif What do you want to call these images within CellProfiler? Red Type the text that one type of image has in common (for TEXT options), or their position in each group (for ORDER option): s1_w3.tif What do you want to call these images within CellProfiler? Green Type the text that one type of image has in common (for TEXT options), or their position in each group (for ORDER option): / What do you want to call these images within CellProfiler? / If using ORDER, how many images are there in each group (i.e. each field of view)? 3 What type of files are you loading? individual images Analyze all subfolders within the selected folder? No Enter the path name to the folder where the images to be loaded are located. Type period (.) for default image folder. D: Note - If the movies contain more than just one image type (e.g., brightfield, fluorescent, field-of-view), add the GroupMovieFrames module. D: Module #2: RescaleIntensity revision - 2 What did you call the image to be rescaled? Blue What do you want to call the rescaled image? Blue2 Rescaling method. (S) Stretch the image (0 to 1). (E) Enter the minimum and maximum values in the boxes below. (G) rescale so all pixels are equal to or Greater than one. (M) Match the maximum of one image to the maximum of another. (C) Convert to 8 bit. See the help for details. Enter min/max below (Method E only): Enter the intensity from the original image that should be set to the lowest value in the rescaled image, or type AA to calculate the lowest intensity automatically from all of the images to be analyzed and AE to calculate the lowest intensity from each image independently. 0 (Method E only): Enter the intensity from the original image that should be set to the highest value in the rescaled image, or type AA to calculate the highest intensity automatically from all of the images to be analyzed and AE to calculate the highest intensity from each image independently. 0.0625 (Method E only): What should the lowest intensity of the rescaled image be (range [0,1])? 0 (Method E only): What should the highest intensity of the rescaled image be (range [0,1])? 1 (Method M only): What did you call the image whose maximum you want the rescaled image to match? Blue Module #3: Resize revision - 1 What did you call the image to be resized? Blue2 What do you want to call the resized image? Blue3 To shrink the image, enter the resizing factor (0 to 1). To enlarge the image, enter the resizing factor (greater than 1). .5 Alternately, leave the shrinking factor set to 1 and enter the desired resulting size in pixels: height,width. This may change the aspect ratio of the image. 100,100 Enter the interpolation method Nearest Neighbor Module #4: RescaleIntensity revision - 2 What did you call the image to be rescaled? Red What do you want to call the rescaled image? Red2 Rescaling method. (S) Stretch the image (0 to 1). (E) Enter the minimum and maximum values in the boxes below. (G) rescale so all pixels are equal to or Greater than one. (M) Match the maximum of one image to the maximum of another. (C) Convert to 8 bit. See the help for details. Enter min/max below (Method E only): Enter the intensity from the original image that should be set to the lowest value in the rescaled image, or type AA to calculate the lowest intensity automatically from all of the images to be analyzed and AE to calculate the lowest intensity from each image independently. 0 (Method E only): Enter the intensity from the original image that should be set to the highest value in the rescaled image, or type AA to calculate the highest intensity automatically from all of the images to be analyzed and AE to calculate the highest intensity from each image independently. 0.0625 (Method E only): What should the lowest intensity of the rescaled image be (range [0,1])? 0 (Method E only): What should the highest intensity of the rescaled image be (range [0,1])? 1 (Method M only): What did you call the image whose maximum you want the rescaled image to match? Blue Module #5: Resize revision - 1 What did you call the image to be resized? Red2 What do you want to call the resized image? Red3 To shrink the image, enter the resizing factor (0 to 1). To enlarge the image, enter the resizing factor (greater than 1). .5 Alternately, leave the shrinking factor set to 1 and enter the desired resulting size in pixels: height,width. This may change the aspect ratio of the image. 100,100 Enter the interpolation method Nearest Neighbor Module #6: RescaleIntensity revision - 2 What did you call the image to be rescaled? Green What do you want to call the rescaled image? Green2 Rescaling method. (S) Stretch the image (0 to 1). (E) Enter the minimum and maximum values in the boxes below. (G) rescale so all pixels are equal to or Greater than one. (M) Match the maximum of one image to the maximum of another. (C) Convert to 8 bit. See the help for details. Enter min/max below (Method E only): Enter the intensity from the original image that should be set to the lowest value in the rescaled image, or type AA to calculate the lowest intensity automatically from all of the images to be analyzed and AE to calculate the lowest intensity from each image independently. 0 (Method E only): Enter the intensity from the original image that should be set to the highest value in the rescaled image, or type AA to calculate the highest intensity automatically from all of the images to be analyzed and AE to calculate the highest intensity from each image independently. 0.0625 (Method E only): What should the lowest intensity of the rescaled image be (range [0,1])? 0 (Method E only): What should the highest intensity of the rescaled image be (range [0,1])? 1 (Method M only): What did you call the image whose maximum you want the rescaled image to match? Blue Module #7: Resize revision - 1 What did you call the image to be resized? Green2 What do you want to call the resized image? Green3 To shrink the image, enter the resizing factor (0 to 1). To enlarge the image, enter the resizing factor (greater than 1). .5 Alternately, leave the shrinking factor set to 1 and enter the desired resulting size in pixels: height,width. This may change the aspect ratio of the image. 100,100 Enter the interpolation method Nearest Neighbor Module #8: IdentifyPrimAutomatic revision - 12 What did you call the images you want to process? Blue3 What do you want to call the objects identified by this module? Nuclei Typical diameter of objects, in pixel units (Min,Max): 5, 40 Discard objects outside the diameter range? Yes Try to merge too small objects with nearby larger objects? No Discard objects touching the border of the image? Yes Select an automatic thresholding method or enter an absolute threshold in the range [0,1]. To choose a binary image, select "Other" and type its name. Choosing ''All'' will use the Otsu Global method to calculate a single threshold for the entire image group. The other methods calculate a threshold for each image individually. "Set interactively" will allow you to manually adjust the threshold during the first cycle to determine what will work well. Otsu Adaptive Threshold correction factor 1 Lower and upper bounds on threshold, in the range [0,1] 0,1 For MoG thresholding, what is the approximate fraction of image covered by objects? 10 Method to distinguish clumped objects (see help for details): Intensity Method to draw dividing lines between clumped objects (see help for details): Intensity Size of smoothing filter, in pixel units (if you are distinguishing between clumped objects). Enter 0 for low resolution images with small objects (~< 5 pixel diameter) to prevent any image smoothing. Automatic Suppress local maxima within this distance, (a positive integer, in pixel units) (if you are distinguishing between clumped objects) Automatic Speed up by using lower-resolution image to find local maxima? (if you are distinguishing between clumped objects) Yes Enter the following information, separated by commas, if you would like to use the Laplacian of Gaussian method for identifying objects instead of using the above settings: Size of neighborhood(height,width),Sigma,Minimum Area,Size for Wiener Filter(height,width),Threshold / What do you want to call the outlines of the identified objects (optional)? Do not save Do you want to fill holes in identified objects? Yes Do you want to run in test mode where each method for distinguishing clumped objects is compared? No Module #9: IdentifyPrimAutomatic revision - 12 What did you call the images you want to process? Red3 What do you want to call the objects identified by this module? H3P Typical diameter of objects, in pixel units (Min,Max): 5,30 Discard objects outside the diameter range? Yes Try to merge too small objects with nearby larger objects? No Discard objects touching the border of the image? Yes Select an automatic thresholding method or enter an absolute threshold in the range [0,1]. To choose a binary image, select "Other" and type its name. Choosing ''All'' will use the Otsu Global method to calculate a single threshold for the entire image group. The other methods calculate a threshold for each image individually. "Set interactively" will allow you to manually adjust the threshold during the first cycle to determine what will work well. Otsu Global Threshold correction factor 1 Lower and upper bounds on threshold, in the range [0,1] 0.10,1 For MoG thresholding, what is the approximate fraction of image covered by objects? 10 Method to distinguish clumped objects (see help for details): Intensity Method to draw dividing lines between clumped objects (see help for details): Intensity Size of smoothing filter, in pixel units (if you are distinguishing between clumped objects). Enter 0 for low resolution images with small objects (~< 5 pixel diameter) to prevent any image smoothing. Automatic Suppress local maxima within this distance, (a positive integer, in pixel units) (if you are distinguishing between clumped objects) Automatic Speed up by using lower-resolution image to find local maxima? (if you are distinguishing between clumped objects) Yes Enter the following information, separated by commas, if you would like to use the Laplacian of Gaussian method for identifying objects instead of using the above settings: Size of neighborhood(height,width),Sigma,Minimum Area,Size for Wiener Filter(height,width),Threshold / What do you want to call the outlines of the identified objects (optional)? Do not save Do you want to fill holes in identified objects? Yes Do you want to run in test mode where each method for distinguishing clumped objects is compared? No Module #10: IdentifySecondary revision - 3 What did you call the primary objects you want to create secondary objects around? H3P What do you want to call the objects identified by this module? H3P_Cells Select the method to identify the secondary objects (Distance - B uses background; Distance - N does not): Distance - N What did you call the images to be used to find the edges of the secondary objects? For DISTANCE - N, this will not affect object identification, only the final display. Red3 Select an automatic thresholding method or enter an absolute threshold in the range [0,1]. To choose a binary image, select "Other" and type its name. Choosing ''All'' will use the Otsu Global method to calculate a single threshold for the entire image group. The other methods calculate a threshold for each image individually. Set interactively will allow you to manually adjust the threshold during the first cycle to determine what will work well. Otsu Adaptive Threshold correction factor 1 Lower and upper bounds on threshold, in the range [0,1] 0,1 For MoG thresholding, what is the approximate fraction of image covered by objects? 10 For DISTANCE, enter the number of pixels by which to expand the primary objects [Positive integer] 15 For PROPAGATION, enter the regularization factor (0 to infinity). Larger=distance,0=intensity 0.05 What do you want to call the outlines of the identified objects (optional)? Do not save Do you want to run in test mode where each method for identifying secondary objects is compared? No Module #11: IdentifyPrimAutomatic revision - 12 What did you call the images you want to process? Green3 What do you want to call the objects identified by this module? Cnndots Typical diameter of objects, in pixel units (Min,Max): 1.2, 5 Discard objects outside the diameter range? Yes Try to merge too small objects with nearby larger objects? No Discard objects touching the border of the image? Yes Select an automatic thresholding method or enter an absolute threshold in the range [0,1]. To choose a binary image, select "Other" and type its name. Choosing ''All'' will use the Otsu Global method to calculate a single threshold for the entire image group. The other methods calculate a threshold for each image individually. "Set interactively" will allow you to manually adjust the threshold during the first cycle to determine what will work well. Otsu Adaptive Threshold correction factor 2 Lower and upper bounds on threshold, in the range [0,1] 0.19,1 For MoG thresholding, what is the approximate fraction of image covered by objects? 10 Method to distinguish clumped objects (see help for details): Intensity Method to draw dividing lines between clumped objects (see help for details): Intensity Size of smoothing filter, in pixel units (if you are distinguishing between clumped objects). Enter 0 for low resolution images with small objects (~< 5 pixel diameter) to prevent any image smoothing. 0 Suppress local maxima within this distance, (a positive integer, in pixel units) (if you are distinguishing between clumped objects) Automatic Speed up by using lower-resolution image to find local maxima? (if you are distinguishing between clumped objects) Yes Enter the following information, separated by commas, if you would like to use the Laplacian of Gaussian method for identifying objects instead of using the above settings: Size of neighborhood(height,width),Sigma,Minimum Area,Size for Wiener Filter(height,width),Threshold / What do you want to call the outlines of the identified objects (optional)? Do not save Do you want to fill holes in identified objects? No Do you want to run in test mode where each method for distinguishing clumped objects is compared? No Module #12: MeasureObjectAreaShape revision - 3 What did you call the objects that you want to measure? Cnndots Do not use Do not use Do not use Do not use Do not use Do not use Would you like to calculate the Zernike features for each object (with lots of objects, this can be very slow)? No Module #13: FilterByObjectMeasurement revision - 4 Which object would you like to filter by, or if using a Ratio, what is the numerator object? Cnndots What do you want to call the filtered objects? Cnndots2 Which category of measurements would you want to filter by? AreaShape Which feature do you want to use? (Enter the feature number - see help for details) 7 For INTENSITY or TEXTURE features, which image''s measurements do you want to use (for other measurements, this will only affect the display)? Green3 Minimum value required: 1.1 Maximum value allowed: 3.2 What do you want to call the outlines of the identified objects (optional)? Do not save Module #14: MeasureObjectIntensity revision - 2 What did you call the greyscale images you want to measure? Green3 What did you call the objects that you want to measure? Cnndots2 Do not use Do not use Do not use Do not use Do not use Module #15: FilterByObjectMeasurement revision - 4 Which object would you like to filter by, or if using a Ratio, what is the numerator object? Cnndots2 What do you want to call the filtered objects? Cnndots3 Which category of measurements would you want to filter by? Intensity Which feature do you want to use? (Enter the feature number - see help for details) 5 For INTENSITY or TEXTURE features, which image''s measurements do you want to use (for other measurements, this will only affect the display)? Green3 Minimum value required: 0.25 Maximum value allowed: 1 What do you want to call the outlines of the identified objects (optional)? Do not save Module #16: Relate revision - 2 What objects are the children objects (subobjects)? Nuclei What are the parent objects? Nuclei What other object do you want to find distances to? (Must be one object per parent object, e.g. Nuclei) None Module #17: Relate revision - 2 What objects are the children objects (subobjects)? Nuclei What are the parent objects? Nuclei What other object do you want to find distances to? (Must be one object per parent object, e.g. Nuclei) None Module #18: Relate revision - 2 What objects are the children objects (subobjects)? Nuclei What are the parent objects? Nuclei What other object do you want to find distances to? (Must be one object per parent object, e.g. Nuclei) None Module #19: SpeedUpCellProfiler revision - 5 Output files should be saved every Nth cycle (1,2,3,...). Note: the output file is always saved after the first or last cycle is processed, no matter what is entered here. 5 Do you want to clear the memory (may not be compatible with some modules)? Yes If yes, which images would you like to remain in memory? Yes Do not use Do not use Do not use Do not use Do not use Do not use Do not use Do not use Do not use Do not use Do not use