An alternative flow cytometry strategy for peripheral blood dendritic cell enumeration in the setting of repetitive GM-CSF dosing.

Journal of Translational Medicine
Kehui WangEdward L Nelson

Abstract

Enumeration of circulating peripheral blood dendritic cells (DCs) is complicated by the absence of a unique cell surface marker expressed on all DC subsets and by the use of various biological adjuvants to modulate the DC compartment, including granulocyte macrophage colony stimulating factor (GM-CSF). Common methods employ a cocktail of antibodies, typically including anti-CD14, to define a lineage negative, MHC class II positive, putative DC population. Reported flow cytometry protocols include highly variable gating strategies and DC identification criteria. Increasing appreciation of DC pleiomorphism, GM-CSF biology, and recognition of CD14 expression in some DC subsets led us to consider an alternative lineage cocktail to improve identification of the circulating DC pool. Standard whole blood staining with appropriate fluorochrome conjugated antibodies to MHC class II and either standard CD14 containing, or an alternate CD66acde containing, lineage cocktail was performed on samples obtained from normal donors and breast cancer patients before and after administration of dose-dense, cytotoxic chemotherapy with daily GM-CSF hematopoetic growth factor support. Putative DCs were enumerated by standard flow cytometry. Data set ...Continue Reading

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Datasets Mentioned

BETA
GM-CSF

Methods Mentioned

BETA
flow cytometry
biopsies
activated cell sorting
fluorescent
FACS
applicable to whole blood flow

Software Mentioned

FlowJo
Graph Pad
SAS
Graph Pad Prism
Excel

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