Support for FCS 3.2?

[alefrol638]

Dear CytoML Team,

I am trying to import gates from a workspace, which works with FCS files from the new BD software FACS CHORUS. The FCS files are FCS 3.2 and the function gives me the error, that the format is wrong:

 gs  <- flowjo_to_gatingset(open_flowjo_xml(wsp, sample_names_from="sampleNode"), name = 1,mc.cores = 7,path = <pathToFile>) #flowWorkspace package

Error: This does not seem to be a valid FCS2.0, FCS3.0 or FCS3.1 file

Is there a way to work with this workspace/FCS files?

Thanks!

[mikejiang]

We don't have official support for 3.2 yet, did relax the check in flowcore https://github.com/RGLab/flowCore/pull/263 so that it can be load there, I can do a similar change in cytolib to allow your workspace parsing proceed, but no guarantee it will work if your fcs is not fully compatible with 3.1

[alefrol638]

all right, I would give it a try

[mikejiang]

https://github.com/RGLab/cytolib/pull/58

[mikejiang]

I pushed the cytolib change to bioconductor develop branch: To trigger the rebuild for CytoML, I bumped its version to 2.15.3 as well

[schraivo]

Hi CytoML Team, I have the same problem as alefrol638. I now use cytolib_2.15.3 and CytoML_2.15.2, should that still throw the same error when loading the FCS3.2 file? Here are the packages I have installed, which should be the latest ones:

*cytolib 2.15.3 2024-08-28 [1] Github (RGLab/cytolib@4f42d85) CytoML 2.15.2 2024-08-22 [1] Github (RGLab/CytoML@b1c491e) flowCore 2.15.3 2024-08-20 [1] Github (RGLab/flowCore@761a60f) flowWorkspace 4.15.5 2024-06-24 [1] Github (RGLab/flowWorkspace@3951e19)**

Thank you!

[mikejiang]

shouldn't throw error, post your sessioninfo and reproducible script/example file so that we can trouble shoot

[schraivo]

Sure, here's an example. You need two files (sorry, didn't have smaller ones available, about 30 MB each): WSP: https://oc.embl.de/index.php/s/g4Yh2gI3lX1cq46 FCS: https://oc.embl.de/index.php/s/qP8i3zpZp9rq0Ho

library(CytoML)
library(flowCore)
library(flowWorkspace)
library(tidyverse)

#Input files
dir <- "path to parent directory of the two downloaded files"
wsp <- list.files(dir, pattern = ".wsp")
fcs <- list.files(dir, pattern = ".fcs")

fc <- as_tibble(exprs(read.FCS(paste0(dir, fcs), truncate_max_range = FALSE)))

#Warning: The flowCore package does not fully support FCS3.2 yet

gs  <- flowjo_to_gatingset(open_flowjo_xml(paste0(dir, wsp), name = 1, sample_names_from = "sampleNode"), name = 1)

#Error: This does not seem to be a valid FCS2.0, FCS3.0 or FCS3.1 file

And the session info.

sessionInfo()
R version 4.3.2 (2023-10-31)
Platform: x86_64-pc-linux-gnu (64-bit)
Running under: Rocky Linux 8.8 (Green Obsidian)

Matrix products: default
BLAS/LAPACK: FlexiBLAS OPENBLAS;  LAPACK version 3.11.0

locale:
 [1] LC_CTYPE=en_US.UTF-8       LC_NUMERIC=C               LC_TIME=en_US.UTF-8        LC_COLLATE=en_US.UTF-8    
 [5] LC_MONETARY=en_US.UTF-8    LC_MESSAGES=en_US.UTF-8    LC_PAPER=en_US.UTF-8       LC_NAME=C                 
 [9] LC_ADDRESS=C               LC_TELEPHONE=C             LC_MEASUREMENT=en_US.UTF-8 LC_IDENTIFICATION=C       

time zone: Europe/Berlin
tzcode source: system (glibc)

attached base packages:
[1] stats     graphics  grDevices utils     datasets  methods   base     

other attached packages:
 [1] lubridate_1.9.3      forcats_1.0.0        stringr_1.5.0        dplyr_1.1.4          purrr_1.0.2         
 [6] readr_2.1.5          tidyr_1.3.1          tibble_3.2.1         ggplot2_3.5.1        tidyverse_2.0.0     
[11] flowWorkspace_4.15.5 flowCore_2.15.3      CytoML_2.15.2       

loaded via a namespace (and not attached):
 [1] gtable_0.3.5        xfun_0.41           Biobase_2.62.0      lattice_0.22-6      tzdb_0.4.0         
 [6] vctrs_0.6.5         tools_4.3.2         generics_0.1.3      stats4_4.3.2        curl_5.1.0         
[11] fansi_1.0.6         pkgconfig_2.0.3     data.table_1.15.4   RColorBrewer_1.1-3  S4Vectors_0.40.2   
[16] graph_1.80.0        lifecycle_1.0.4     compiler_4.3.2      munsell_0.5.1       htmltools_0.5.7    
[21] yaml_2.3.8          hexbin_1.28.3       pillar_1.9.0        RProtoBufLib_2.14.1 tidyselect_1.2.1   
[26] digest_0.6.33       stringi_1.7.12      fastmap_1.1.1       grid_4.3.2          colorspace_2.1-0   
[31] cli_3.6.3           magrittr_2.0.3      ncdfFlow_2.48.0     RBGL_1.78.0         XML_3.99-0.16.1    
[36] utf8_1.2.4          withr_3.0.0         scales_1.3.0        timechange_0.3.0    rmarkdown_2.25     
[41] httr_1.4.7          matrixStats_1.3.0   gridExtra_2.3       cytolib_2.15.3      hms_1.1.3          
[46] evaluate_0.23       knitr_1.45          rlang_1.1.4         Rcpp_1.0.12         glue_1.7.0         
[51] Rgraphviz_2.46.0    BiocGenerics_0.48.1 rstudioapi_0.15.0   jsonlite_1.8.8      R6_2.5.1           
[56] plyr_1.8.9          zlibbioc_1.48.2     ggcyto_1.30.2  

Thank you!

[mikejiang]

The links provided above seem to be identical and point to the same fcs file, where is wsp file?

[mikejiang]

I think I know the reason,cytolib is statically linked to flowWorkspace/CytoML packages, looks like you have cytolib up to date, try to re-install/compile flowWorkspace/CytoML so that they also contain the latest change from cytolib I tested and it worked for me

library(flowWorkspace)

file="opensource/D118_2dg.fcs"
load_cytoframe_from_fcs(file)

[schraivo]

Hi Mike. Please apologize the delayed response. Unfortunately, it didn't work for me. I updated the links in my original post, hoping that you could have another look. Thank you very much!

[mikejiang]

I was able to reproduce your error with fcs file you provided last time, then i re compiled/build packages in question i mentioned above, and problem went away. so it's definitely something out of sync in your current installation. i can't further help if it is your installation environment issue, try to switch to another machine or person to verify. alternatively, purge your R and install everything from scratch

[schraivo]

Sounds good, thank you, Mike!