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Question: opencyto Error in .validity_check_alias
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gravatar for biomiha
14 months ago by
biomiha10
UK/Cambridge
biomiha10 wrote:

Hi,

 

I'm trying to use the OpenCyto platform to streamline our FACS analysis. I've generated a GatingSet from a FlowJo .acs archive - incidentally for other users who've experienced similar problems with .wsp files, I've found that if the workspace is bundled together into an archive then parseWorkspace seems to work. 

I then generated a template file using the templateGen function. After modifying the gating method column in the csv template file I try to import the template back using the gatingTemplate function, whereby I get an

    Error in .validity_check_alias(this_row[, alias]) : 
    Q4: CD4-FITC-H- , CD8-PE-H-contains illegal character: |,&,:,/

error. 

I think the error is self descriptive, i.e. it's because of the colon character but it would seem to me that

a) quite many FlowJo workspaces will have some sort of quadrant gate, which will have this type of syntax

b) if I rename the aliases in the template I get

Error in .cpp_setNodeName(x@pointer, sampleNames(x), y, value) : 
  Q1: CD4-FITC-H- , CD8-PE-H+ not found!

 

Is there a workaround for this?

Also, I'm struggling somewhat to figure out what gating methods to use in the template file. Is there any documentation on this? That would be much appreciated. 

Many thanks.

Miha

ADD COMMENTlink modified 14 months ago by Jiang, Mike1.2k • written 14 months ago by biomiha10

Found this (https://www.bioconductor.org/packages/devel/bioc/vignettes/openCyto/inst/doc/HowToAutoGating.html) so that will be a great place to start re gating methods. Still curious as to how to solve the syntax error.

ADD REPLYlink written 14 months ago by biomiha10
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gravatar for Jiang, Mike
14 months ago by
Jiang, Mike1.2k
(Private Address)
Jiang, Mike1.2k wrote:

Can you post your template? Also the traceback output for the error after you renamed the alias

ADD COMMENTlink written 14 months ago by Jiang, Mike1.2k

For some reason Dropbox has changed their sharing options and I had to add an email to share the template. I hope it was delivered to the right email address.

ADD REPLYlink written 14 months ago by biomiha10
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gravatar for Jiang, Mike
14 months ago by
Jiang, Mike1.2k
(Private Address)
Jiang, Mike1.2k wrote:

In your parent column, you still referred to the old alias name 'Q1: CD4-FITC-H- , CD8-PE-H+', which no longer exists in your template.

You want to keep these updated.

Also, in your pop column, you should have simple '+' or '-' symbols instead of the full channel/marker names, which contain these symbols in themselves and will confuse the template parser on the quadrant pattern. Read the manual carefully http://bioconductor.org/packages/3.5/bioc/vignettes/openCyto/inst/doc/HowToWriteCSVTemplate.html especially the red notes.

We have enforced this rule in the latest devel version https://github.com/RGLab/openCyto/issues/156

Lastly, I don't think you should use flowClust for all of your quradrant gates. Please follow these example csv templates shipped with the package to do the things in the right way

https://github.com/RGLab/openCyto/tree/trunk/inst/extdata/gating_template

ADD COMMENTlink written 14 months ago by Jiang, Mike1.2k

Hi Mike,

Thanks for your reply and comments. I'll read through the documentation you suggested - I have started collating all the different vignettes, tutorials and templates I can find scattered in various places on the internet.

Re your template comment: I just used the templateGen function on an existing GatingHierarchy object. I understand from your links that the "pop" column needs to be updated but it was in fact generated by the templateGen function, so perhaps that would need updating as well? My initial template file (i.e. template1.csv) included the full alias names and then because that generated a syntax error I replaced the Q1: (Q2:, Q3: and Q4:) in the hope of generating a valid template file - in this case template2.csv.

And, yes, I completely agree that using flowClust for all the quradrant gates is wrong, but to be honest I hadn't even come to the gating methods and just pasted something in the hope of generating a valid template file. Understanding the numerous gating methods and gating arguments is next on my to do list.

Many thanks for this.

Miha

ADD REPLYlink written 14 months ago by biomiha10

Incidentally, do you still have the gt_080.csv template file anywhere? I'm trying to follow the example here but the link from the tutorials (https://www.dropbox.com/s/jdigjss903q3sgu/gt_080.csv) is broken. Or is that just ICS_080.csv now instead?

ADD REPLYlink modified 14 months ago • written 14 months ago by biomiha10
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