WebImageProcessingUsageandHelpThe interface consists of twodrop-downmenusdenoted as "Data" and "Jobs".Allinput, intermediate and final results are accessible via the "Data"menuwhile allcomputationsareaccessibleviathe"Jobs"menu.Thefollowingareguidelinesforacommonworkflowofsteps.

Images/TilesUpload

1. GotoData>Tilescollections

2. Clickon"Createnewcollection"3. Giveitauniquenameandclickon“Save”

4. Uploadtheimagesusingeither:o the"Addfiles/folder"buttonso thedraganddroptarget

AfteruploadingtheimagesandgoingbacktoTilesCollection,thisiswhatyourpageshouldlooklike:

Note: File upload canbe slow.Once files areuploaded theyhave tobe convertedinto ome.tif format.We are usingOMEto supportmetadata storage and retrievalalongtheentirepipelineusingawelldefinedstandardformat.

Fileconversiontoome.tifmaybeslowerthantheupload.

MaskLabelling

1. GotoJobs>Masklabellingjobs2. Clickon"Createnewjob"3. Giveitauniquejobname4. Specifytheimagecollection5. Specifytheconnectedness(fourorheight)

Thisstepisusedtolabelmaskimages(suchastheonesgeneratedbysegmentationsjobs)forusebyfeatureextractionsoftware.

Stitching

1. GotoJobs>Stitchingjobs2. Clickon"Createnewjob"3. Giveitauniquejobname4. Specifywhichimagecollectiontouse5. Setuptheparameters

o MISTo Stagemetadata

Mustwaitfortilestobeuploadedandconvertedtoome.tifbeforelaunching.Jobnameisusedlatertospecifywhichstitchingvectortouse.Needtolaunch1stitchingpercollection.

Segmentation

1. GotoJobs>Segmentationjobs2. Clickon"Createnewjob"3. Giveitauniquejobname4. Specifytheimagecollectiontosegment5. SpecifyEGTparameters

Only required if youwant to flat-field correct images and view the segmentationmaskinthefinalvisualization.Segmentationisdoneperimagecollection.Segmentation job will create a new tile collection to hold the results.Collectionwillbenamedusingthesegmentationjobname.

FlatFieldCorrection

1. GotoJobs>Flatfieldcorrectionjobs2. Clickon"Createnewjob"3. Giveitauniquejobname4. SpecifytheRawImages5. SpecifytheSegmentedImages

If you want to flat-field correct any image collection you must have previouslygenerated segmentation masks for that collection. Job will create a new tilecollectiontoholdtheresults.If thetilesnamesoftheselectedrawandsegmentedtilescollectiondonotmatch,youwillneedtoprovidethepatternsofeachcollection.Thesystemwilltrytoguessthepatternsautomaticallybutyoushoulddoublecheck.

IntensityScaling

1. GotoJobs>Intensityscalingjobs2. Clickon"Createnewjob"3. Giveitauniquejobname4. Specifytheimagecollection5. Specifyre-scalingtype.Usetruncationunlessthereisacompellingreasonnot

to.

Job will create a new tile collection to hold the results.Webbrowsersonlysupportviewing8bppimages. Ifno intensityrescaling isdonethesystemwillmaptheentire16bitrange[0-65535]intothe8bitrange[0-255].Each image collection you wish to view must be rescaled independently.Therearetwotypesofintensityscaling:

Truncation(preferred)By default will saturate the bottom and top 1% of intensities, linearlymapping intensities into that range. This is the same as Fiji/ImageJ auto-contrast.If range start and end are specified those values are used instead ofcomputingrangestartas1stpercentileandendas99thpercentile.

GammaCorrectionThisperformsanon-linear(exponential)rescaling.Bydefaultstartandendare1stpercentileand99thpercentile.

PyramidBuilding

1. GotoJobs>Pyramidjobs2. Clickon"Createnewjob"3. Giveitauniquejobname4. Specifytheimagecollection5. Specifythestitchingvector.Thestitchingvectormusthavebeengeneratedby

apreviouslycompletedstitchingjob.

Before images can be viewed, individual image tiles need to be converted intopyramidsthatcanbeviewedinthebrowser.Iftilesnamesintilescollectionandstitchingvectordiffer,youmustspecifythetilespattern. The system will try to guess the patterns automatically but you shoulddoublecheck.

DeepZoomVisualization

1. GotoData>Visualization2. Clickon"Createnewvisualization"3. Giveitthevisualizationauniquename4. Atthispointcreateoneormoregroups5. Givethegroupalabelandclick"+"6. Addlayers(pyramids)tothegroupwith"+".(Forexample,addTransmitted,

Excitation,Segmentation(ifpyramidwasbuiltfromthemasks))

This stepmust be executed once all pyramids have beenbuilt.Multiple pyramidscanbecombinedintoasinglevisualizationwithmultiplelayers.

ImageTessellation

1. GotoJobs>Tessellationjobs2. Clickon"Createnewjob"3. Giveitauniquejobname4. Specifythemaskwidthofthefinaltessellationimage5. Specifythemaskheightofthefinaltessellationimage6. Specifytheshapeinatessellation(squareorhexagon)7. Specifytheradiusoftheshapeinatessellation

o Forsquareshapes,theradiusisonehalfofitswidth.o Forhexagonal shapes, the radius is thedistancebetween the center

andthesummit.

This step is independentof all other steps. It is executed togenerateamask sub-dividing a segmentation result. Tessellation masks are used together withsegmentationmasks tocomputespatially local image features for studyingspatialheterogeneity.

ImageAssembling

1. GotoJobs>ImageAssemblingjobs2. Clickon"Createnewjob"3. Giveitauniquejobname4. Specifythestitchingvector5. Specifytheimagetilecollection

Thisstepissimilartothepyramidbuildingstepbutinsteadofbuildingapyramid,itbuilds big images. Those big images can then be used to compute features usingthefeatureextractionsoftware.

ExampleWorkflow

Thefigurebelowillustratepossibleworkflowsofcomputationalstepsthathavetobeexecutedinordertovisuallyinspecttwoimagecollectionsofoverlappingfieldsof views (FOV). The web image processing always includes stitching, pyramidbuilding,andvisualizationcreation.Iftheimageshavepixeldepthmorethan8bitsperpixel(BPP)thenintensityrescalinghastobeexecuted.Flatfieldcorrectionandsegmentation steps are optional. However, they are important for quantitativeanalyses.

ExampleofaworkflowusingtheWebImageProcessingsystemtoprocesstwoimagecollections(transmittedandexcitationchannels)acquiredatthesame

time(thechannelsareregistered).