Staging (phenoimager2mc), max-projections

[kbestak]

Hi, this is a PR for our branch of MCMICRO where we've added a staging step before illumination specific for unmixed single-tile outputs from Akoya's InForm software as part of the push to make MCMICRO compatible with SignalStar, a method for multicycle multispectral imaging.

In addition, I've added a max-projection functionality to recyze as described below in the change points - max projection of nuclear markers is returned as channel 0 and max projection of membrane markers is returned as channel 1. This functionality would mostly benefit Cellpose segmentation in my opinion.

• Add staging step before illumination - start-at: staging
  • inputs should be saved in staging folder, outputs will be stored in raw and continue as a typical mcmicro run.
• phenoimager2mc added as a staging module - staging-method: phenoimager2mc
• illumination is run by default if start-at: staging is given and can be toggled off with illumination: false
• Add max projection option to recyze on multiple nuclear and/or membrane channels provided - output will have nuclear channel 0 and membrane channel 1
  • segmentation-max-projection: true
  • segmentation-channel: 1 3 8 9
  • segmentation-nuclear-channel: 1 8
  • segmentation-membrane-channel: 3 9

Example parameter file for running staging, registration, segmentation with cellpose using the above options and quantification:

workflow:
  start-at: staging
  stop-at: quantification
  illumination: false
  staging-method: phenoimager2mc
  segmentation-recyze: true
  segmentation-max-projection: true
  segmentation-channel: 1 5 7 8 11
  segmentation-nuclear-channel: 5 11
  segmentation-membrane-channel: 1 7 8
  segmentation: cellpose
options:
  phenoimager2mc: -m 6 --normalization max
  ashlar: --align-channel 4 --flip-y
  cellpose: --pretrained_model cyto --chan 1 --chan2 0 --no_npy

Let me know if this is of interest and if changes would have to be made,

Best,
Krešo

[ArtemSokolov]

Thanks for submitting this, @kbestak.

Looks like CI test are failing with No subdirectories found in staging directory.
I think you may need to wrap

stagingDirs = Channel.fromPath("${params.in}/staging/*", type: 'dir')
    .ifEmpty { error "No subdirectories found in staging directory" }

inside an if/else:

// Look for staging directories only if the user is starting from the staging step
stagingDirs = (wfp['start-at'] == 'staging') ?
    Channel.fromPath("${params.in}/staging/*", type: 'dir')
        .ifEmpty { error "No subdirectories found in staging directory" } : Channel.empty()

I had to do similar thing for all other steps, so I defined a function here: https://github.com/labsyspharm/mcmicro/blob/master/main.nf#L44-L50

[kbestak]

Thank you for highlighting this, I've addressed the issue, added validation for the max projection / nuclear-membrane specifications, excluded the mentioned staging parameter (now start-at: staging is enough for staging to run), and caught a bug with providing recyze with the max-projection tag that was fixed now.