Experiment Overview

Repository ID: FR-FCM-Z2YR Experiment name: HIMC control replicates-Gates MIFlowCyt score: 24.00%
Primary researcher: Mike Leipold PI/manager: Holden Maecker Uploaded by: Mike Leipold
Experiment dates: 2018-01-25 - 2018-05-17 Dataset uploaded: Apr 2019 Last updated: Apr 2019
Keywords: [PBMC] [healthy controls] [human] [mass cytometry] [CyTOF] [replicates] Manuscripts:
Organizations: None
Purpose: PBMCs from a previously characterized single healthy control donor were included in barcoded mass cytometry (CyTOF) samples as QC/QA controls in the study of other vaccination timecourse samples. The reagents for each plate were combined fresh each day. However, all reagents were from the same commercial lot or same in-house conjugation prep for the entire study. For each plate, a cryopreserved vial of the healthy control donor PBMCs was thawed, then spiked into a barcoded sample as a QC/QA measure of staining procedure and sample running on the machine.
Conclusion: The healthy control files were not used further in the analysis of the study samples. However, the general CyTOF community may find use for these 59 normalized debarcoded healthy control replicates from multiple plates run across approximately 4 months. In particular, we can see an opportunity for others to use these files in the development of batch correction algorithms.
Comments: 22 Plates were run. For each plate, replicates subaliquots of the same healthy control PBMCs were spiked into separate barcoded samples. The samples were stained (fresh cocktails and other reagent dilutions each day), then Fluidigm EQ normalization beads were added and the samples run on a Helios mass cytometer. For each plate, the resulting raw FCS files were normalized using the Premessa R package (Finck et al style normalization), using the Beads files from Plate 1 as a reference. The normalized files were then debarcoded using the barcoding key and the Premessa R package. For each sample, the healthy control PBMC sample was used as a QC/QA measure in the analysis of the other donor samples studied. For this upload, the healthy control FCS files were renamed more clearly using keywords in Mac FlowJo X and re-exported as newly named FCS files. No cell events were removed.
Funding: This work was funded by Center for Human Systems Immunology at Stanford by the Bill and Melinda Gates Foundation OPP1113682. We would also like to thank our collaborators at in the Dr. Pollard lab in the Oxford Vaccine Group for their panel suggestions and other donor samples. We would like to thank labs of Dr. Catherine Blish and Dr. Eugene Butcher at Stanford University for their advice on antibody clone selection for markers related to NK cells and to chemokine receptors, respectively.
Quality control: None
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