cytoqc for lyoplate

Load gs

panel <- "tcell"
gslist <- sapply(centers, function(center) {
  message("Center: ", center)
  gs <- load_gs(file.path(path, center, panel))
  gs_clone(gs)
})

QC Check gates

cqc_data <- cqc_gs_list(gslist)

#group by gates
groups <- cqc_group(cqc_data, "gate")
su <- summary(groups)
knit_print(su)

group_id	nObject	gate
2	5	4- 8+, 4- 8+/38- DR-, 4- 8+/38- DR+, 4- 8+/38+ DR-, 4- 8+/38+ DR+, 4- 8+/CCR7- 45RA-, 4- 8+/CCR7- 45RA+, 4- 8+/CCR7+ 45RA-, 4- 8+/CCR7+ 45RA+, 4+ 8-, 4+ 8-/38- DR-, 4+ 8-/38- DR+, 4+ 8-/38+ DR-, 4+ 8-/38+ DR+, 4+ 8-/CCR7- 45RA-, 4+ 8-/CCR7- 45RA+, 4+ 8-/CCR7+ 45RA-, 4+ 8-/CCR7+ 45RA+, CD3, DNT, DPT, LYM, not dead, root, singlets
1	2	4- 8+, 4- 8+/38- DR-, 4- 8+/38- DR+, 4- 8+/38+ DR-, 4- 8+/38+ DR+, 4- 8+/CCR7- 45RA-, 4- 8+/CCR7- 45RA+, 4- 8+/CCR7+ 45RA-, 4- 8+/CCR7+ 45RA+, 4+ 8-, 4+ 8-/38- DR-, 4+ 8-/38- DR+, 4+ 8-/38+ DR-, 4+ 8-/38+ DR+, 4+ 8-/CCR7- 45RA-, 4+ 8-/CCR7- 45RA+, 4+ 8-/CCR7+ 45RA-, 4+ 8-/CCR7+ 45RA+, CD3, DNT, DPT, LYM, not dead, root

knit_print(diff(su))

group_id	nObject	gate
2	5	singlets

#vis the difference
plot_diff(groups)

# match reference
match_result <- cqc_match_reference(groups, ref = 1, select = c(2))
match_result

group_id	Not in reference	Missing channels
2	singlets

solution <- cqc_find_solution(match_result)
solution

Proposed change
singlets

cqc_fix(solution)

summary(cqc_group(cqc_data, "gate"))

group_id	nObject	gate
1	7	4- 8+, 4- 8+/38- DR-, 4- 8+/38- DR+, 4- 8+/38+ DR-, 4- 8+/38+ DR+, 4- 8+/CCR7- 45RA-, 4- 8+/CCR7- 45RA+, 4- 8+/CCR7+ 45RA-, 4- 8+/CCR7+ 45RA+, 4+ 8-, 4+ 8-/38- DR-, 4+ 8-/38- DR+, 4+ 8-/38+ DR-, 4+ 8-/38+ DR+, 4+ 8-/CCR7- 45RA-, 4+ 8-/CCR7- 45RA+, 4+ 8-/CCR7+ 45RA-, 4+ 8-/CCR7+ 45RA+, CD3, DNT, DPT, LYM, not dead, root

QC check for channel

groups <- cqc_group(cqc_data, "channel")
su <- summary(groups)
knit_print(su)

group_id	nObject	channel
1	1	<Alexa Fluor 488-A>, <AmCyan-A>, <APC-A>, <APC-Cy7-A>, <Pacific Blue-A>, <PE YG-A>, <PE-Cy7 YG-A>, <PerCP-Cy5-5-A>, FSC-A, SSC-A, Time
2	1	<Am Cyan-A>, <APC-A>, <APC-Cy7-A>, <FITC-A>, <Pacific Blue-A>, <PE-A>, <PE-Cy7-A>, <PerCP-Cy5-5-A>, FSC-A, FSC-H, FSC-W, SSC-A, SSC-H, SSC-W, Time
3	1	<AmCyan-A>, <APC-A>, <APC-Cy7-A>, <FITC-A>, <Pacific Blue-A>, <PE Cy7 YG-A>, <PE-A>, <PerCP-Cy5-5-A>, FSC-A, FSC-W, SSC-A, SSC-W, Time
4	1	<APC-A>, <APC-Cy7-A>, <FITC-A>, <PE-A>, <PE-Cy7-A>, <PerCP-Cy5-5-A>, <V450-A>, <V500-A>, FSC-A, SSC-A, Time
5	1	<APC-A>, <APC-H7-A>, <BD Horizon V450-A>, <BD Horizon V500-A>, <FITC-A>, <PE-A>, <PE-Cy7-A>, <PerCP-Cy5-5-A>, FSC-A, FSC-H, FSC-W, SSC-A, SSC-H, SSC-W, Time
6	1	<APC-A>, <APC-H7-A>, <FITC-A>, <PE-A>, <PE-Cy7-A>, <PerCP-Cy5-5-A>, <V450-A>, <V500-A>, FSC-A, FSC-H, SSC-A, SSC-H, Time
7	1	<B515-A>, <B710-A>, <G560-A>, <G780-A>, <R660-A>, <R780-A>, <V450-A>, <V545-A>, FSC-A, FSC-H, FSC-W, SSC-A, SSC-H, SSC-W, Time

diff(su)

group_id	nObject	channel
1	1	<Alexa Fluor 488-A>, <AmCyan-A>, <APC-A>, <APC-Cy7-A>, <Pacific Blue-A>, <PE YG-A>, <PE-Cy7 YG-A>, <PerCP-Cy5-5-A>
2	1	<Am Cyan-A>, <APC-A>, <APC-Cy7-A>, <FITC-A>, <Pacific Blue-A>, <PE-A>, <PE-Cy7-A>, <PerCP-Cy5-5-A>, FSC-H, FSC-W, SSC-H, SSC-W
3	1	<AmCyan-A>, <APC-A>, <APC-Cy7-A>, <FITC-A>, <Pacific Blue-A>, <PE Cy7 YG-A>, <PE-A>, <PerCP-Cy5-5-A>, FSC-W, SSC-W
4	1	<APC-A>, <APC-Cy7-A>, <FITC-A>, <PE-A>, <PE-Cy7-A>, <PerCP-Cy5-5-A>, <V450-A>, <V500-A>
5	1	<APC-A>, <APC-H7-A>, <BD Horizon V450-A>, <BD Horizon V500-A>, <FITC-A>, <PE-A>, <PE-Cy7-A>, <PerCP-Cy5-5-A>, FSC-H, FSC-W, SSC-H, SSC-W
6	1	<APC-A>, <APC-H7-A>, <FITC-A>, <PE-A>, <PE-Cy7-A>, <PerCP-Cy5-5-A>, <V450-A>, <V500-A>, FSC-H, SSC-H
7	1	<B515-A>, <B710-A>, <G560-A>, <G780-A>, <R660-A>, <R780-A>, <V450-A>, <V545-A>, FSC-H, FSC-W, SSC-H, SSC-W

Channels are very different across centers so move on to check marker

#summarise marker groups
groups <- cqc_group(cqc_data, "marker")
su <- summary(groups)
knit_print(su)

group_id	nObject	marker
6	2	CD197, CD3, CD38, CD4, CD45RA, CD8, HLA-DR, Live Green
1	1	CCR7 PE, CD3 V450, CD38 APC, CD4 PerCP-Cy55, CD45RA PE-Cy7, CD8 APC-H7, HLA-DR V500, Live Dead FITC
2	1	CCR7, CD3, CD38, CD4, CD45RA, CD8, HLA DR, Live/Dead
3	1	CCR7, CD3, CD38, CD4, CD45RA, CD8, HLA-DR, LIVE
4	1	CCR7, CD3, CD38, CD4, CD45RA, CD8, HLADR, LIVE_GREEN
5	1	CD197, CD3, CD38, CD4, CD45RA, CD8, HLA-DR, LIVE DEAD

knit_print(diff(su))

group_id	nObject	marker
6	2	CD197, CD3, CD38, CD4, CD45RA, CD8, HLA-DR, Live Green
1	1	CCR7 PE, CD3 V450, CD38 APC, CD4 PerCP-Cy55, CD45RA PE-Cy7, CD8 APC-H7, HLA-DR V500, Live Dead FITC
2	1	CCR7, CD3, CD38, CD4, CD45RA, CD8, HLA DR, Live/Dead
3	1	CCR7, CD3, CD38, CD4, CD45RA, CD8, HLA-DR, LIVE
4	1	CCR7, CD3, CD38, CD4, CD45RA, CD8, HLADR, LIVE_GREEN
5	1	CD197, CD3, CD38, CD4, CD45RA, CD8, HLA-DR, LIVE DEAD

Markers are more standardized and go ahead to further clean it

res <- cqc_match_reference(groups, ref = 3, select = c(1,2,4,5,6)) 
res

group_id	Not in reference	Missing channels
1	CCR7 PE,CD3 V450,CD38 APC,CD4 PerCP-Cy55,CD45RA PE-Cy7,CD8 APC-H7,HLA-DR V500,Live Dead FITC	CCR7,CD3,CD38,CD4,CD45RA,CD8,HLA-DR,LIVE
2	HLA DR,Live/Dead	HLA-DR,LIVE
4	HLADR,LIVE_GREEN	HLA-DR,LIVE
5	CD197,LIVE DEAD	CCR7,LIVE
6	CD197,Live Green	CCR7,LIVE

solution <- cqc_find_solution(res, max.distance = 0.6)
solution

Proposed change
CCR7 PE –> CCR7
CD38 APC –> CD38
CD3 V450 –> CD3
HLA-DR V500 –> HLA-DR
CD45RA PE-Cy7 –> CD45RA
HLA DR –> HLA-DR
Live/Dead –> LIVE
HLADR –> HLA-DR
LIVE_GREEN –> LIVE
CD197 –> CCR7
LIVE DEAD –> LIVE
Live Green –> LIVE

cqc_fix(solution)

update checks

groups <- cqc_group(cqc_data, "marker")
knit_print(summary(groups))

group_id	nObject	marker
2	6	CCR7, CD3, CD38, CD4, CD45RA, CD8, HLA-DR, LIVE
1	1	CCR7, CD3, CD38, CD4 PerCP-Cy55, CD45RA, CD8 APC-H7, HLA-DR, Live Dead FITC

Relax string matching to clean up the rest

res <- cqc_match_reference(groups, ref = 2, select = c(1)) 
res

group_id	Not in reference	Missing channels
1	CD4 PerCP-Cy55,CD8 APC-H7,Live Dead FITC	CD4,CD8,LIVE

solution <- cqc_find_solution(res, max.distance = 0.8)
solution

Proposed change
CD8 APC-H7 –> CD8
Live Dead FITC –> LIVE
CD4 PerCP-Cy55 –> CD4

cqc_fix(solution)

groups <- cqc_group(cqc_data, "marker")
knit_print(summary(groups))

group_id	nObject	marker
1	7	CCR7, CD3, CD38, CD4, CD45RA, CD8, HLA-DR, LIVE

Use the marker as reference to standardize the channels across centers

panel <- cf_get_panel(gs_cyto_data(cqc_data[[1]])[[1]], skip_na = TRUE)
panel

## # A tibble: 8 x 2
##   channel         marker  
##   <I<chr>>        <I<chr>>
## 1 <APC-A>         CD38    
## 2 <APC-H7-A>      CD8     
## 3 <FITC-A>        LIVE    
## 4 <PerCP-Cy5-5-A> CD4     
## 5 <V450-A>        CD3     
## 6 <V500-A>        HLA-DR  
## 7 <PE-A>          CCR7    
## 8 <PE-Cy7-A>      CD45RA

cqc_set_panel(cqc_data, panel, ref.col = "marker")
groups <- cqc_group(cqc_data, "panel")
knit_print(summary(groups))

channel	marker	group_id	nObject
<APC-A>	CD38	1	7
<APC-H7-A>	CD8
<FITC-A>	LIVE
<PE-A>	CCR7
<PE-Cy7-A>	CD45RA
<PerCP-Cy5-5-A>	CD4
<V450-A>	CD3
<V500-A>	HLA-DR

Refresh QC report

groups <- cqc_group(cqc_data, "channel")
su <- summary(groups)
knit_print(su)

group_id	nObject	channel
1	3	<APC-A>, <APC-H7-A>, <FITC-A>, <PE-A>, <PE-Cy7-A>, <PerCP-Cy5-5-A>, <V450-A>, <V500-A>, FSC-A, FSC-H, FSC-W, SSC-A, SSC-H, SSC-W, Time
4	2	<APC-A>, <APC-H7-A>, <FITC-A>, <PE-A>, <PE-Cy7-A>, <PerCP-Cy5-5-A>, <V450-A>, <V500-A>, FSC-A, SSC-A, Time
2	1	<APC-A>, <APC-H7-A>, <FITC-A>, <PE-A>, <PE-Cy7-A>, <PerCP-Cy5-5-A>, <V450-A>, <V500-A>, FSC-A, FSC-H, SSC-A, SSC-H, Time
3	1	<APC-A>, <APC-H7-A>, <FITC-A>, <PE-A>, <PE-Cy7-A>, <PerCP-Cy5-5-A>, <V450-A>, <V500-A>, FSC-A, FSC-W, SSC-A, SSC-W, Time

knit_print(diff(su))

group_id	nObject	channel
1	3	FSC-H, FSC-W, SSC-H, SSC-W
2	1	FSC-H, SSC-H
3	1	FSC-W, SSC-W

Remove `H/W` channels

res <- cqc_match_reference(groups, ref = 4, select = c(1,2,3)) 
res

group_id	Not in reference	Missing channels
1	FSC-H,FSC-W,SSC-H,SSC-W
2	FSC-H,SSC-H
3	FSC-W,SSC-W

solution <- cqc_find_solution(res, max.distance = 0.6)
solution

Proposed change
FSC-H
FSC-W
SSC-H
SSC-W

cqc_fix(solution)

Coerce it directly into single `GatingSet` (zero-copying)

gs <- merge_list_to_gs(cqc_data)
gs

cytoqc for lyoplate

QC for tcell panel

Load gs

QC Check gates

QC check for channel

Channels are very different across centers so move on to check marker

Markers are more standardized and go ahead to further clean it

update checks

Relax string matching to clean up the rest

Use the marker as reference to standardize the channels across centers

Refresh QC report

Remove H/W channels

Coerce it directly into single GatingSet (zero-copying)

Remove `H/W` channels

Coerce it directly into single `GatingSet` (zero-copying)