Unfamiliar error codes when initiating tSNE analysis in cytofkit

[theeclaire]

When setting up a new tSNE analysis using cytofkit I keep getting the following error message which seems to refer to issues with my .fcs files. The arg# changes with different attempts, but the analysis stalls each time I try to run all files. I have trailled running with smaller numbers of samples, which works, and standardising all file names/removing typos, which does not improve performance. Any advice on how to get the analsysis up and running would be much appreciated.

---

Extract expression data... Error in (function (..., deparse.level = 1) : number of columns of matrices must match (see arg 116)

[raman91]

Hi @theeclaire, have you tested with the example data ? It looks like issue with the fcs files. Do let us know if more assistance is required. Thanks

[theeclaire]

Hi @raman91. Thanks for the suggestion, yes I have and works with smaller .fcs file sets but the error code when I run the full set gives me no clue what is going wrong with the files; what does it mean by: "number of columns of matrices must match (see arg 31)" - is there somewhere I can fing out what 'arg 31' is?

All .fcs files (n>1500) were processed and exported identically so I am not clear why some samples would not be suitable for the program or how to troubleshoot the mismatched 'column number' - any ideas?

[theeclaire]

(n.b. the arg number changes each time, hence why the arg# differs between the two comments (i.e. 113 and 31)

[raman91]

Hi @theeclaire, did you provide metadata in your analysis? Please make sure that all uploaded fcs files are included in the metadata. Can you pls check and confirm? Thanks

[theeclaire]

Thanks @raman91. I am a new R user and not sure what you mean by inlcuding the metadata; here is some more data on how the files were generated which I hope addresses your query. I generated the files for the analysis in flowjo by applying compensation and minimal gating (leukocytes, singlets) identically across samples and exporting the equivalent nodes with these settings applied. I then used the GUI to select the exported files for the analysis. I’ve narrowed the issue down to 4 samples and seems to be a mismatch in columns but it is not clear to me from the error code exactly what the issue is with these samples relative to the others since they were treated the same. This is also not apparent when I go back to flowjo and investigate the raw vs exported files, so I am assuming that the issue is with the R processing of the files; hence trying to work out what the error codes are referring to. Many thanks.