I am trying to modify the compensation matrix of an fcs file using flowCore.

I have no error or warning but it seems that the modified compensation was not applied to the FlowFrame object.

Below is a code example.

Can anyone help me?

library(FlowCore)
frame <- read.FCS(myFile)    # this is an fcs 3.0 file
#let's create a dummy matrix for the example
mat <- diag(10)
mat[1,2] <- 0.5

# change the name for channels to match the names in the fcs 3.0 file
 colnames(mat) <- c("FL1-A",  "FL2-A",  "FL3-A",  "FL4-A",  "FL5-A",  "FL6-A",  "FL7-A",  "FL8-A",  "FL9-A",  "FL10-A")
 rownames(mat) <- c("FL1-A",  "FL2-A",  "FL3-A",  "FL4-A",  "FL5-A",  "FL6-A",  "FL7-A",  "FL8-A",  "FL9-A",  "FL10-A")

#These names match the names from the fcs file
> colnames(frame)
 [1] "FS-A"   "FS-W"   "SS-H"   "SS-A"   "SS-W"   "FL1-A"  "FL2-A"  "FL3-A"  "FL4-A"  "FL5-A"  "FL6-A"  "FL7-A"  "FL8-A" 
[14] "FL9-A"  "FL10-A" "TIME"

# Apply compensation to the flowFrame
  comp <- compensation(mat)
  frame_comp <- compensate(frame, comp)

# if we check if any modification of the original compensation matrix have been done
spillover(fcs)$`$SPILLOVER` == spillover(frame_comp)$`$SPILLOVER`

>         6    7    8    9   10   11   12   13   14   15
 [1,] TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE
 [2,] TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE
 [3,] TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE
 [4,] TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE
 [5,] TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE
 [6,] TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE
 [7,] TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE
 [8,] TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE
 [9,] TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE
[10,] TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE TRUE

# So no modification in the compensation have been done by the compensate() function..

sessionInfo( )

R version 4.2.1 (2022-06-23 ucrt)
Platform: x86_64-w64-mingw32/x64 (64-bit)
Running under: Windows 10 x64 (build 19044)

Matrix products: default

locale:
[1] LC_COLLATE=French_France.utf8  LC_CTYPE=French_France.utf8    LC_MONETARY=French_France.utf8
[4] LC_NUMERIC=C                   LC_TIME=French_France.utf8    

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

other attached packages:
[1] XML_3.99-0.12  flowCore_2.8.0

loaded via a namespace (and not attached):
 [1] compiler_4.2.1      RProtoBufLib_2.8.0  cytolib_2.8.0       tools_4.2.1         Rcpp_1.0.9         
 [6] Biobase_2.56.0      S4Vectors_0.34.0    BiocGenerics_0.42.0 matrixStats_0.62.0  RcppParallel_5.1.5 
[11] stats4_4.2.1

Eventually I don't know why the previous code didn't work to modify the compensation matrix (I guess that my understanding of the compensate() function is insufficient).

But I did found a workaround. Adding the following codeline was all that was needed to modify the compensation matrix.

keyword(frame)$`$SPILLOVER` <- mat