## Loading required package: flowCore
## Loading required package: lattice

We change the way w parameter is estimated in estimateLogicle function

Regular flow data

marker  <- 'CD3'
chnl <- chnls[1]

data distribution on CD3

data <- exprs(fr[, chnl])
den <- density(data, adjust = 2)
plot(den, main = paste("raw flow", marker, sep = ":"))

  • high value at 4.9653072 decade
  • neg value at -3.6467819 decade

Estimated logicle (default)

trans <- estimateLogicle(fr, channels = chnl)
sapply(c("t","m", "a", "w"), function(param)as.numeric(format(as.vector(environment(trans@transforms[[chnl]]@f)[[param]]), digits = 2)))
##         t         m         a         w 
## 261590.00      4.50      0.00      0.47
fr_trans <- transform(fr, trans)
data <- exprs(fr_trans[, chnl])
den <- density(data, adjust = 2)
g <- mindensity(fr_trans, chnl)
plot(den, main = paste("transformed flow", marker, sep = ":"))
abline(v = g@min, col = "red")

xyplot(`SSC-A`~`CD3`, fr_trans, smooth = F, xbin = 64, filter = g)

Estimated logicle (method = “peak”)

trans <- estimateLogicle(fr, channels = chnl, method = "peak", plot = T)

sapply(c("t","m", "a", "w"), function(param)as.numeric(format(as.vector(environment(trans@transforms[[chnl]]@f)[[param]]), digits = 2)))
##        t        m        a        w 
## 261590.0      4.5      0.0      1.1
fr_trans <- transform(fr, trans)
data <- exprs(fr_trans[, chnl])
den <- density(data, adjust = 2)
g <- mindensity(fr_trans, chnl)
plot(den, main = paste("transformed flow", marker, sep = ":"))
abline(v = g@min, col = "red")

xyplot(`SSC-A`~`CD3`, fr_trans, smooth = F, xbin = 64, filter = g)

marker  <- 'IFNg'
chnl <- chnls[2]

IFNg

data <- exprs(fr[, chnl])
den <- density(data, adjust = 2)
plot(den, main = paste("raw flow", marker,sep = ":"))

Estimated logicle (default)

trans <- estimateLogicle(fr, channels = chnl)
sapply(c("t","m", "a", "w"), function(param)as.numeric(format(as.vector(environment(trans@transforms[[chnl]]@f)[[param]]), digits = 2)))
##         t         m         a         w 
## 261590.00      4.50      0.00      0.62
fr_trans <- transform(fr, trans)
data <- exprs(fr_trans[, chnl])
den <- density(data, adjust = 2)
g <- tailgate(fr_trans, chnl)
plot(den, main = paste("transformed flow", marker,sep = ":"))
abline(v = g@min, col = "red")

xyplot(`SSC-A`~`IFNg`, fr_trans, smooth = F, xbin = 64, filter = g)

Estimated logicle (method = “peak”)

trans <- estimateLogicle(fr, channels = chnl, method = "peak", plot = T)

sapply(c("t","m", "a", "w"), function(param)as.numeric(format(as.vector(environment(trans@transforms[[chnl]]@f)[[param]]), digits = 2)))
##         t         m         a         w 
## 261590.00      4.50      0.00      0.97
fr_trans <- transform(fr, trans)
data <- exprs(fr_trans[, chnl])
den <- density(data, adjust = 2)
g <- tailgate(fr_trans, chnl)
plot(den, main = paste("transformed flow", marker,sep = ":"))
abline(v = g@min, col = "red")

xyplot(`SSC-A`~`IFNg`, fr_trans, smooth = F, xbin = 64, filter = g)

CyTOF data

## the text section does not end with delimiter: \\. The last keyword is dropped.
## the text section does not end with delimiter: \\. The last keyword is dropped.
marker  <- 'CD3'
chnl <- chnls[1]

CD3

data <- exprs(fr[, chnl])
den <- density(data, adjust = 2)
plot(den, main = paste("raw cytof", marker,sep = ":"))

  • high value at 4.4361783 decade
  • low at -0.0623533 decade

Estimated logicle (default)

trans <- estimateLogicle(fr, channels = chnl)
sapply(c("t","m", "a", "w"), function(param)as.numeric(format(as.vector(environment(trans@transforms[[chnl]]@f)[[param]]), digits = 2)))
##        t        m        a        w 
## 2.73e+04 4.50e+00 0.00e+00 4.80e-02
fr_trans <- transform(fr, trans)
data <- exprs(fr_trans[, chnl])
den <- density(data, adjust = 2)
g <- mindensity(fr_trans, chnl)
plot(den, main = paste("transformed cytof", marker,sep = ":"))
abline(v = g@min, col = "red")

xyplot(`Cell_length`~`(Cd112)Dd`, fr_trans, smooth = F, xbin = 64, filter = g, checkName = F)

Estimated logicle (method = “peak”)

trans <- estimateLogicle(fr, channels = chnl, method = "peak", plot = T)

sapply(c("t","m", "a", "w"), function(param)as.numeric(format(as.vector(environment(trans@transforms[[chnl]]@f)[[param]]), digits = 2)))
##       t       m       a       w 
## 27300.0     4.5     0.0     1.3
fr_trans <- transform(fr, trans)
data <- exprs(fr_trans[, chnl])
den <- density(data, adjust = 2)
g <- mindensity(fr_trans, chnl)
plot(den, main = paste("transformed cytof", marker,sep = ":"))
abline(v = g@min, col = "red")

xyplot(`Cell_length`~`(Cd112)Dd`, fr_trans, smooth = F, xbin = 64, filter = g, checkName = F)

marker <- 'IFNg'
chnl <- chnls[2]

data distribution on IFNg

data <- exprs(fr[, chnl])
den <- density(data, adjust = 2)
plot(den, main = paste("raw cytof", marker,sep = ":"))

  • high at 3.4233612 decade
  • low at -0.0635091 decade

Estimated logicle (default)

trans <- estimateLogicle(fr, channels = chnl)
sapply(c("t","m", "a", "w"), function(param)as.numeric(format(as.vector(environment(trans@transforms[[chnl]]@f)[[param]]), digits = 2)))
##       t       m       a       w 
## 2650.00    4.50    0.00    0.55
fr_trans <- transform(fr, trans)
data <- exprs(fr_trans[, chnl])
den <- density(data, adjust = 2)
g <- tailgate(fr_trans, chnl)
plot(den, main = paste("transformed cytof", marker,sep = ":"))
abline(v = g@min, col = "red")

xyplot(`Cell_length`~`(Yb170)Dd`, fr_trans, smooth = F, xbin = 64, filter = g, checkName = F)

Estimated logicle (method = “peak”)

trans <- estimateLogicle(fr, channels = chnl, method = "peak", plot = T)

sapply(c("t","m", "a", "w"), function(param)as.numeric(format(as.vector(environment(trans@transforms[[chnl]]@f)[[param]]), digits = 2)))
##      t      m      a      w 
## 2650.0    4.5    0.0    1.5
fr_trans <- transform(fr, trans)
data <- exprs(fr_trans[, chnl])
den <- density(data, adjust = 2)
g <- tailgate(fr_trans, chnl)
plot(den, main = paste("transformed cytof", marker,sep = ":"))
abline(v = g@min, col = "red")

xyplot(`Cell_length`~`(Yb170)Dd`, fr_trans, smooth = F, xbin = 64, filter = g, checkName = F)