Hi, This is Wilson. I am trying to write a loop function to possibly mass analyse a set of flow frame files. However my sequence does not seem to recognise the "Subset" function utilised by the flowCore package. The loop terminates after the recognition of the Pico matrix (highlighted portion).

Pls kindly take a look and advise if I wrote any part wrong.

Thanks

Wilson

library(flowCore)

df1<-list.files("G://Trial/") ##directory with list of files to be read in  

setwd("G://Trial/")

for(i in df1){

xN <- as.character(i)

y <- read.FCS(xN, transformation="linearize", which.lines=NULL,
              alter.names=FALSE, column.pattern=NULL, invert.pattern = FALSE,
              decades=0, ncdf = FALSE, min.limit=NULL, dataset=NULL, emptyValue=TRUE)  

logTrans <- logTransform(transformationId="log10-transformation", logbase=10, r=1, d=1)
trans <- transformList(c("SSC-Log_Height", "FSC-Log_Height", "FL1-Log_Height", "FL2-Log_Height", "FL3-Log_Height", "FL4-Log_Height", "FL5-Log_Height", "FL6-Log_Height", "FL7-Log_Height" , "FL8-Log_Height", "FL9-Log_Height", "FL10-Log_Height", "FL11-Log_Height"), logTrans)

xNtrans <- transform(y, trans) 

##Matrix to be applied for data extraction 

Pico <- matrix(c(1,1,5,5,0,2.5,4.85,0),ncol=2,nrow=4) ## (y-axis, x-axis)

colnames(Pico) <- c("FL4-Log_Height","FL2-Log_Height") 
pico <- polygonGate(filterId="Counts", .gate = Pico)

xNs<-Subset(xN,pico)

Rect <- matrix(c(1,5,5,1,1,1,5,5),ncol=2,nrow=4) ## (y-axis, x-axis)

colnames(Rect) <- c("FL4-Log_Height","SSC-Log_Height")
rect <- polygonGate(filterId="Counts", .gate = Rect)

xNF<-Subset(xNs,rect)

z<-exprs(xNF) } 

lapply(names(z), function(x){ write.table(z, file = paste("G://Trial/", x,".txt", sep = ""),sep="\t",row.names=F,col.names=T,quote=F)})

It would be easier to diagnose if you gave the exact error message, but as a guess, flowcore::Subset requires the first argument be a flow object (such as your xNtrans ) whereas you're feeding it xN which is a character.

It is recommended to use `add` method to append these gates to the `GatingSet` object from flowWorkspace package to capture your entire gating process for better organizing, archiving and retrieving the gating hierarchy. See the vignette of flowCore for more details.