NF1_analysis_4channel.cppipe

CellProfiler Pipeline: http://www.cellprofiler.org
Version:5
DateRevision:424
GitHash:
ModuleCount:17
HasImagePlaneDetails:False

Images:[module_num:1|svn_version:'Unknown'|variable_revision_number:2|show_window:False|notes:['Images module is left blank since we are giving the path to the images in the CLI command']|batch_state:array([], dtype=uint8)|enabled:True|wants_pause:False]
    :
    Filter images?:Images only
    Select the rule criteria:and (extension does isimage) (directory doesnot containregexp "[\\\\/]\\.")

Metadata:[module_num:2|svn_version:'Unknown'|variable_revision_number:6|show_window:False|notes:['Metadata is extracted from the file and folder names using regular expressions.']|batch_state:array([], dtype=uint8)|enabled:True|wants_pause:False]
    Extract metadata?:Yes
    Metadata data type:Text
    Metadata types:{"Channel": "integer", "FileLocation": "text", "Frame": "text", "Plate": "text", "Series": "text", "Site": "integer", "Stain": "float", "Well": "text"}
    Extraction method count:2
    Metadata extraction method:Extract from file/folder names
    Metadata source:File name
    Regular expression to extract from file name:(?P<Well>[A-Z]{1}[0-9]{1,2})_01_(?P<Channel>[1-4]{1})_(?P<Site>[0-9]{1,2})_(?P<Stain>DAPI|CY5|GFP|RFP)
    Regular expression to extract from folder name:(?P<Date>[0-9]{4}_[0-9]{2}_[0-9]{2})$
    Extract metadata from:All images
    Select the filtering criteria:and (file does contain "")
    Metadata file location:Elsewhere...|
    Match file and image metadata:[]
    Use case insensitive matching?:No
    Metadata file name:None
    Does cached metadata exist?:No
    Metadata extraction method:Extract from file/folder names
    Metadata source:Folder name
    Regular expression to extract from file name:^(?P<Plate>.*)_(?P<Well>[A-P][0-9]{2})_s(?P<Site>[0-9])_w(?P<ChannelNumber>[0-9])
    Regular expression to extract from folder name:Corrected_(?P<Plate>Plate_3_prime|Plate_[0-9]{1})
    Extract metadata from:All images
    Select the filtering criteria:and (file does contain "")
    Metadata file location:Elsewhere...|
    Match file and image metadata:[]
    Use case insensitive matching?:No
    Metadata file name:None
    Does cached metadata exist?:No

NamesAndTypes:[module_num:3|svn_version:'Unknown'|variable_revision_number:8|show_window:False|notes:['This pipeline assigns names to 4 channels specifically:', '', 'DAPI', 'GFP', 'CY5', 'RFP']|batch_state:array([], dtype=uint8)|enabled:True|wants_pause:False]
    Assign a name to:Images matching rules
    Select the image type:Grayscale image
    Name to assign these images:DNA
    Match metadata:[]
    Image set matching method:Order
    Set intensity range from:Image metadata
    Assignments count:4
    Single images count:0
    Maximum intensity:255.0
    Process as 3D?:No
    Relative pixel spacing in X:1.0
    Relative pixel spacing in Y:1.0
    Relative pixel spacing in Z:1.0
    Select the rule criteria:and (file does contain "DAPI")
    Name to assign these images:DAPI
    Name to assign these objects:Cell
    Select the image type:Grayscale image
    Set intensity range from:Image metadata
    Maximum intensity:255.0
    Select the rule criteria:and (file does contain "GFP")
    Name to assign these images:GFP
    Name to assign these objects:Nucleus
    Select the image type:Grayscale image
    Set intensity range from:Image metadata
    Maximum intensity:255.0
    Select the rule criteria:and (file does contain "RFP")
    Name to assign these images:RFP
    Name to assign these objects:Cytoplasm
    Select the image type:Grayscale image
    Set intensity range from:Image metadata
    Maximum intensity:255.0
    Select the rule criteria:and (file does contain "CY5")
    Name to assign these images:CY5
    Name to assign these objects:Speckle
    Select the image type:Grayscale image
    Set intensity range from:Image metadata
    Maximum intensity:255.0

Groups:[module_num:4|svn_version:'Unknown'|variable_revision_number:2|show_window:False|notes:['We do not use the Groups module.']|batch_state:array([], dtype=uint8)|enabled:True|wants_pause:False]
    Do you want to group your images?:No
    grouping metadata count:1
    Metadata category:None

IdentifyPrimaryObjects:[module_num:5|svn_version:'Unknown'|variable_revision_number:15|show_window:False|notes:['The parameters are the same as the analysis pipeline for 3 channels.']|batch_state:array([], dtype=uint8)|enabled:True|wants_pause:False]
    Select the input image:DAPI
    Name the primary objects to be identified:Nuclei
    Typical diameter of objects, in pixel units (Min,Max):30,90
    Discard objects outside the diameter range?:Yes
    Discard objects touching the border of the image?:Yes
    Method to distinguish clumped objects:None
    Method to draw dividing lines between clumped objects:Shape
    Size of smoothing filter:10
    Suppress local maxima that are closer than this minimum allowed distance:7.0
    Speed up by using lower-resolution image to find local maxima?:Yes
    Fill holes in identified objects?:After both thresholding and declumping
    Automatically calculate size of smoothing filter for declumping?:Yes
    Automatically calculate minimum allowed distance between local maxima?:Yes
    Handling of objects if excessive number of objects identified:Continue
    Maximum number of objects:500
    Use advanced settings?:Yes
    Threshold setting version:12
    Threshold strategy:Global
    Thresholding method:Otsu
    Threshold smoothing scale:1.3488
    Threshold correction factor:1.0
    Lower and upper bounds on threshold:0.0,1.0
    Manual threshold:0.0
    Select the measurement to threshold with:None
    Two-class or three-class thresholding?:Three classes
    Log transform before thresholding?:No
    Assign pixels in the middle intensity class to the foreground or the background?:Foreground
    Size of adaptive window:50
    Lower outlier fraction:0.05
    Upper outlier fraction:0.05
    Averaging method:Mean
    Variance method:Standard deviation
    # of deviations:2.0
    Thresholding method:Minimum Cross-Entropy

IdentifySecondaryObjects:[module_num:6|svn_version:'Unknown'|variable_revision_number:10|show_window:False|notes:['The parameters are the same as the analysis pipeline for 3 channels.']|batch_state:array([], dtype=uint8)|enabled:True|wants_pause:False]
    Select the input objects:Nuclei
    Name the objects to be identified:Cells
    Select the method to identify the secondary objects:Propagation
    Select the input image:RFP
    Number of pixels by which to expand the primary objects:10
    Regularization factor:0.05
    Discard secondary objects touching the border of the image?:Yes
    Discard the associated primary objects?:No
    Name the new primary objects:Nuclei
    Fill holes in identified objects?:Yes
    Threshold setting version:12
    Threshold strategy:Global
    Thresholding method:Otsu
    Threshold smoothing scale:1.3488
    Threshold correction factor:1.0
    Lower and upper bounds on threshold:0.0,1.0
    Manual threshold:0.0
    Select the measurement to threshold with:None
    Two-class or three-class thresholding?:Three classes
    Log transform before thresholding?:No
    Assign pixels in the middle intensity class to the foreground or the background?:Foreground
    Size of adaptive window:50
    Lower outlier fraction:0.05
    Upper outlier fraction:0.05
    Averaging method:Mean
    Variance method:Standard deviation
    # of deviations:2.0
    Thresholding method:Otsu

IdentifyTertiaryObjects:[module_num:7|svn_version:'Unknown'|variable_revision_number:3|show_window:False|notes:['The parameters are the same as the analysis pipeline for 3 channels.']|batch_state:array([], dtype=uint8)|enabled:True|wants_pause:False]
    Select the larger identified objects:Cells
    Select the smaller identified objects:Nuclei
    Name the tertiary objects to be identified:Cytoplasm
    Shrink smaller object prior to subtraction?:Yes

MeasureColocalization:[module_num:8|svn_version:'Unknown'|variable_revision_number:5|show_window:False|notes:['Measure whole image and within objects ']|batch_state:array([], dtype=uint8)|enabled:True|wants_pause:False]
    Select images to measure:CY5, DAPI, GFP, RFP
    Set threshold as percentage of maximum intensity for the images:15.0
    Select where to measure correlation:Both
    Select objects to measure:Cells, Cytoplasm, Nuclei
    Run all metrics?:Yes
    Calculate correlation and slope metrics?:Yes
    Calculate the Manders coefficients?:Yes
    Calculate the Rank Weighted Colocalization coefficients?:Yes
    Calculate the Overlap coefficients?:Yes
    Calculate the Manders coefficients using Costes auto threshold?:Yes
    Method for Costes thresholding:Faster

MeasureGranularity:[module_num:9|svn_version:'Unknown'|variable_revision_number:4|show_window:False|notes:['Measures within images and objects (when parameter is turned on).']|batch_state:array([], dtype=uint8)|enabled:True|wants_pause:False]
    Select images to measure:CY5, DAPI, GFP, RFP
    Measure within objects?:Yes
    Select objects to measure:Cells, Cytoplasm, Nuclei
    Subsampling factor for granularity measurements:0.25
    Subsampling factor for background reduction:0.25
    Radius of structuring element:10
    Range of the granular spectrum:16

MeasureObjectIntensity:[module_num:10|svn_version:'Unknown'|variable_revision_number:4|show_window:False|notes:['Measures intensity within objects']|batch_state:array([], dtype=uint8)|enabled:True|wants_pause:False]
    Select images to measure:CY5, DAPI, GFP, RFP
    Select objects to measure:Cells, Cytoplasm, Nuclei

MeasureImageIntensity:[module_num:11|svn_version:'Unknown'|variable_revision_number:4|show_window:False|notes:['Measures whole image intensity not aggregated within objects']|batch_state:array([], dtype=uint8)|enabled:True|wants_pause:False]
    Select images to measure:CY5, DAPI, GFP, RFP
    Measure the intensity only from areas enclosed by objects?:No
    Select input object sets:Cells, Cytoplasm, Nuclei
    Calculate custom percentiles:No
    Specify percentiles to measure:10,90

MeasureObjectNeighbors:[module_num:12|svn_version:'Unknown'|variable_revision_number:3|show_window:False|notes:['Measure nuclei neighbors that are adjacent to each other.']|batch_state:array([], dtype=uint8)|enabled:True|wants_pause:False]
    Select objects to measure:Nuclei
    Select neighboring objects to measure:Nuclei
    Method to determine neighbors:Adjacent
    Neighbor distance:50
    Consider objects discarded for touching image border?:Yes
    Retain the image of objects colored by numbers of neighbors?:No
    Name the output image:ObjectNeighborCount
    Select colormap:Blues
    Retain the image of objects colored by percent of touching pixels?:No
    Name the output image:PercentTouching
    Select colormap:Oranges

MeasureObjectNeighbors:[module_num:13|svn_version:'Unknown'|variable_revision_number:3|show_window:False|notes:['Measure cell neighbors that are adjacent to each other.']|batch_state:array([], dtype=uint8)|enabled:True|wants_pause:False]
    Select objects to measure:Cells
    Select neighboring objects to measure:Cells
    Method to determine neighbors:Adjacent
    Neighbor distance:50
    Consider objects discarded for touching image border?:Yes
    Retain the image of objects colored by numbers of neighbors?:No
    Name the output image:ObjectNeighborCount
    Select colormap:Blues
    Retain the image of objects colored by percent of touching pixels?:No
    Name the output image:PercentTouching
    Select colormap:Oranges

MeasureObjectIntensityDistribution:[module_num:14|svn_version:'Unknown'|variable_revision_number:6|show_window:False|notes:['Measure object intensity distributions witin objects using center x,y coords for each single object.', '', "We calculate Zernikes for magnitudes and phases as we are interested in seeing phase even if it doesn't help with classifying phenotypes."]|batch_state:array([], dtype=uint8)|enabled:True|wants_pause:False]
    Select images to measure:CY5, DAPI, GFP, RFP
    Hidden:3
    Hidden:1
    Hidden:0
    Calculate intensity Zernikes?:Magnitudes and phase
    Maximum zernike moment:9
    Select objects to measure:Nuclei
    Object to use as center?:These objects
    Select objects to use as centers:None
    Select objects to measure:Cytoplasm
    Object to use as center?:These objects
    Select objects to use as centers:None
    Select objects to measure:Cells
    Object to use as center?:These objects
    Select objects to use as centers:None
    Scale the bins?:Yes
    Number of bins:4
    Maximum radius:100

MeasureObjectSizeShape:[module_num:15|svn_version:'Unknown'|variable_revision_number:3|show_window:False|notes:['Measure object size and shape.']|batch_state:array([], dtype=uint8)|enabled:True|wants_pause:False]
    Select object sets to measure:Cells, Cytoplasm, Nuclei
    Calculate the Zernike features?:Yes
    Calculate the advanced features?:No

MeasureTexture:[module_num:16|svn_version:'Unknown'|variable_revision_number:7|show_window:False|notes:['Measure texture within objects and the whole images.']|batch_state:array([], dtype=uint8)|enabled:True|wants_pause:False]
    Select images to measure:CY5, DAPI, GFP, RFP
    Select objects to measure:Cells, Cytoplasm, Nuclei
    Enter how many gray levels to measure the texture at:256
    Hidden:1
    Measure whole images or objects?:Both
    Texture scale to measure:3

ExportToDatabase:[module_num:17|svn_version:'Unknown'|variable_revision_number:28|show_window:False|notes:['Output the morphology features into an SQLite database.']|batch_state:array([], dtype=uint8)|enabled:True|wants_pause:False]
    Database type:SQLite
    Database name:DefaultDB
    Add a prefix to table names?:No
    Table prefix:MyExpt_
    Output file location:Default Output Folder|NF1_SchwannCell_data/CellProfiler_pipelines/Analysis_Output/Plate2_Output
    Create a CellProfiler Analyst properties file?:No
    Database host:
    Username:
    Password:
    Name the SQLite database file:nf1_analysis.sqlite
    Calculate the per-image mean values of object measurements?:No
    Calculate the per-image median values of object measurements?:No
    Calculate the per-image standard deviation values of object measurements?:No
    Calculate the per-well mean values of object measurements?:No
    Calculate the per-well median values of object measurements?:No
    Calculate the per-well standard deviation values of object measurements?:No
    Export measurements for all objects to the database?:All
    Select the objects:Cells,Cytoplasm,Nuclei
    Maximum # of characters in a column name:64
    Create one table per object, a single object table or a single object view?:One table per object type
    Enter an image url prepend if you plan to access your files via http:
    Write image thumbnails directly to the database?:No
    Select the images for which you want to save thumbnails:
    Auto-scale thumbnail pixel intensities?:Yes
    Select the plate type:None
    Select the plate metadata:None
    Select the well metadata:None
    Include information for all images, using default values?:Yes
    Properties image group count:1
    Properties group field count:1
    Properties filter field count:0
    Workspace measurement count:1
    Experiment name:NF1_Data
    Which objects should be used for locations?:None
    Enter a phenotype class table name if using the Classifier tool in CellProfiler Analyst:
    Export object relationships?:No
    Overwrite without warning?:Never
    Access CellProfiler Analyst images via URL?:No
    Select the classification type:Object
    Select an image to include:None
    Use the image name for the display?:Yes
    Image name:Channel1
    Channel color:red
    Do you want to add group fields?:No
    Enter the name of the group:
    Enter the per-image columns which define the group, separated by commas:ImageNumber, Image_Metadata_Plate, Image_Metadata_Well
    Do you want to add filter fields?:No
    Automatically create a filter for each plate?:No
    Create a CellProfiler Analyst workspace file?:No
    Select the measurement display tool:ScatterPlot
    Type of measurement to plot on the X-axis:Image
    Enter the object name:None
    Select the X-axis measurement:None
    Select the X-axis index:ImageNumber
    Type of measurement to plot on the Y-axis:Image
    Enter the object name:None
    Select the Y-axis measurement:None
    Select the Y-axis index:ImageNumber