This repository contains code to perform analysis of a surface marker panel flow cytometry experiment from a convalescent cohort of COVID-19 subjects who were either hospitalized (n=20) or not hospitalized (n=40). See the accompanying paper published in JCI Insight on Feb 23, 2021: Comorbid illnesses are associated with altered adaptive immune responses to SARS-CoV-2

Running the analysis

1. Download the FCS files for Study SDY1680 from ImmPort:
   • Click on the Download tab
   • Log into ImmPort
   • Check the ResultFiles box
   • Click Download. Aspera will be launched. Files for ICS and Surface Marker experiments will be downloaded at the same time. Download will take some time (total of 18.3 GB, which took 2 hrs and 42 min to download)
   • These steps for downloading from ImmPort are current as of Feb 8, 2021.
2. Clone or download this repository. The code depends on the current directory structure. Create an R project in this top-level folder. Open it using RStudio.
3. Install dependencies. Analysis was completed using R version 3.6.3 on a computer running Ubuntu 18.04. The code in this branch works with openCyto 1.24.0, CytoML 1.12.1, flowCore 1.52.1, and flowWorkspace 3.34.1. Dependencies are listed in the renv.lock file. If you would like to use renv to install dependencies into this project's renv subfolder, install renv and then run renv::activate() and renv::restore() (see Collaborating with renv). You can install dependencies however way you prefer.
   • TODO: Test renv workflow and upgrade repo with a branch for R 4.0
4. Run scripts/00_20210208_Copy_and_Rename_Surface_Marker_ImmPort_FCS_Files.R after modifying the variable ImmPort_dl_path.
5. Run the rest of the R and Rmd scripts in numerical order. Output will get placed into the out and processed_data subfolders (you may want to copy the current output folders somewhere to have an original copy).