singlecellexperiments markers after clustering
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achaillon • 0
@achaillon-14117
Last seen 23 months ago

Hello I am using the CATALYST::cluster function on my flow data. My panel includes 13 markers

panel$fcs_colname [1] "Alexa Fluor 700-A" "PE-CF594-A" "APC-H7-A" "BV421-A" "BV711-A" "BV786-A" "BV605-A" "PE-Cy7-A" [9] "BV650-A" "APC-A" "PE-A" "PerCP-Cy5-5-A" "Alexa Fluor 488-A"

and are present in my flowset object

all(panel$fcs_colname %in% colnames(fs1)) [1] TRUE

but after running

CATALYST::cluster(sce, features = NULL, xdim = 10, ydim = 10, maxK = 20, seed = 1234)

I only have 11 markers left....

DataFrame with 11 rows and 4 columns channel_name marker_name marker_class used_for_clustering CD3 Alexa Fluor 700-A CD3 type TRUE CD4 PE-CF594-A CD4 type TRUE CD8 APC-H7-A CD8 type TRUE HLA-DR BV421-A HLA-DR type TRUE CD38 BV711-A CD38 state TRUE CCR7 BV786-A CCR7 type TRUE CD45RA BV605-A CD45RA type TRUE PD-1 PE-Cy7-A PD-1 state TRUE TIM-3 BV650-A TIM-3 state TRUE CD73 PerCP-Cy5-5-A CD73 state TRUE CD160 Alexa Fluor 488-A CD160 state TRUE

I am probably missing something obvious... can you help?

thank you!

CATALYST SingleCellExperiment • 486 views
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Hard to read the output above - but if I'm not mistaken, the missing channels are APC-A and PE-A. These are the only ones not containing any numbers. So my guess is this:

  • By default, prepData() will retain only mass channels in the assay data. All none mass channels (e.g. in CyTOF, these are time and event length) are moved to the int_colData().
  • Thus, when you construct the SingleCellExperiment, the object already has only 11 features, and this is not the result of running cluster().
  • Now, since this is FACS data, there are no mass and no-mass channels, and we want to retain all parameters.
  • To enforce this, you should run prepData(..., FACS = TRUE). All 13 markers will then remain in the main assay data.
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I confirm it works with FACS=TRUE. thank you so much. I will follow with another question but using a different thread

thank you

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@crowellh-11823
Last seen 9 months ago
University of Zurich, Switzerland

Solution:

Run prepData() with argument FACS = TRUE.

Explanation:

By default, prepData() will retain only mass channels in the assay data. All none mass channels (e.g. in CyTOF, these are time and event length) are moved to the int_colData(). Since this is FACS data, there are no mass and no-mass channels, and we want to retain all parameters. Setting FACS = TRUE when running prepData() to construct the SingleCellExperiment enforces this.

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