Handling Flow Cytometry Data in R
Cytomics Workshop
April, 2017
© 2015 by Wade Rogers
Handling Flow Cytometry Data: flowCore
• flowCore is a base package that supports reading and manipulation
of FCS data files
• The fundamental object that encapsulates the data in an FCS file is
a flowFrame
• A container object that holds a collection of flowFrames is called a
flowSet
• In the next slides we will go over
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reading an FCS file
gating
compensation
transformation
visualization
Check out the example data
Read an FCS file, summarize the flowFrame
Apply the lymphocyte gate with Subset
needs to be
transformed because
it is rendering the linear data
in the FCS file
hasn’t been compensated!
Fingerprinting Flow Cytometry Data: flowFP
• flowFP
 aims to transform flow cytometric data into a form amenable to
algorithmic analysis tools
 Acts as in intermediate step between acquisition of high-throughput
FCM data and empirical modeling, machine learning and knowledge
discovery
 Implements ideas from
Roederer M, Moore W, Treister A, Hardy RR & Herzenberg LA. Probability binning comparison:
a metric for quantitating multivariate distribution differences. Cytometry 45:47-55, 2001.
and
Rogers WT, Moser AR, Holyst HA, Bantly A, Mohler ER III, Scangas G, and Moore JS, Cytometric
Fingerprinting: Quantitative Characterization of Multivariate Distributions, Cytometry 73A: 430-441,
2008.
The basic idea
• Subdivide multivariate space into bins
 Call this a “model” of the space
• For each flowFrame in a flowSet, count the number of events in each
bin in the model
• Flatten the collection of counts for a flowFrame into a 1D feature
vector
• Combine all of the feature vectors together into a n x m matrix
 n = number of flowFrames (instances)
 m = number of bins in the model (features)
• Also, tag each event with its bin membership
 facilitates visualization, interpretation
 can be used for gating
Probability Binning
Probability Binning
Probability Binning
Probability Binning
Bin Number
> plot (mod, fs)
Class Constructors
• flowFPModel (base class)
 Consumes a flowFrame or flowSet
 Produces a model, which is a recipe for subdividing multivariate space
• flowFP
 Consumes a flowFrame or flowSet, and a flowFPModel
 Produces a flowFP, which represents the multivariate probability density
function as a fingerprint
 Also tags each event with its bin membership
• flowFPPlex
 Consumes a collection of flowFPs
 The flowFPPlex is a container object to facilitate handling large and
complex collections of flowFPs