title: qFlow Cytometry-Based Receptoromic Screening: A High-Throughput Quantification Approach Informing Biomarker Selection and Nanosensor Development.
abstract: Nanosensor-based detection of biomarkers can improve medical diagnosis; however, a critical factor in nanosensor development is deciding which biomarker to target, as most diseases present several biomarkers. Biomarker-targeting decisions can be informed via an understanding of biomarker expression. Currently, immunohistochemistry (IHC) is the accepted standard for profiling biomarker expression. While IHC provides a relative mapping of biomarker expression, it does not provide cell-by-cell readouts of biomarker expression or absolute biomarker quantification. Flow cytometry overcomes both these IHC challenges by offering biomarker expression on a cell-by-cell basis, and when combined with calibration standards, providing quantitation of biomarker concentrations: this is known as qFlow cytometry. Here, we outline the key components for applying qFlow cytometry to detect biomarkers within the angiogenic vascular endothelial growth factor receptor family. The key aspects of the qFlow cytometry methodology include: antibody specificity testing, immunofluorescent cell labeling, saturation analysis, fluorescent microsphere calibration, and quantitative analysis of both ensemble and cell-by-cell data. Together, these methods enable high-throughput quantification of biomarker expression.
authors.affiliation: Bioengineering Department, University of Illinois at Urbana-Champaign, Urbana, IL, USA.
authors.forename: Si
authors.initials: S
authors.lastname: Chen
authors.affiliation: Bioengineering Department, University of Illinois at Urbana-Champaign, Urbana, IL, USA.
authors.forename: Jared
authors.initials: J
authors.lastname: Weddell
authors.affiliation: Chicago College of Osteopathic Medicine, Downers Grove, IL, USA.
authors.forename: Pavan
authors.initials: P
authors.lastname: Gupta
authors.affiliation: Indiana University School of Medicine, Indianapolis, IN, USA.
authors.forename: Grace
authors.initials: G
authors.lastname: Conard
authors.affiliation: Bioengineering Department, California Institute of Technology, Pasadena, CA, USA.
authors.forename: James
authors.initials: J
authors.lastname: Parkin
authors.affiliation: Bioengineering Department, University of Illinois at Urbana-Champaign, Urbana, IL, USA.
authors.forename: Princess I
authors.initials: PI
authors.lastname: Imoukhuede
mesh_terms: D015415:Biomarkers; D015374:Biosensing Techniques / Q000379:methods*; D055808:Drug Discovery / Q000379:methods; D005434:Flow Cytometry / Q000379:methods*; D057166:High-Throughput Screening Assays; D061307:Human Umbilical Vein Endothelial Cells / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D018919:Neovascularization, Physiologic / Q000187:drug effects; D011956:Receptors, Cell Surface / Q000378:metabolism*; D012984:Software; D013223:Statistics as Topic / Q000379:methods
chemical_list: D015415:Biomarkers; D011956:Receptors, Cell Surface
keywords: Angiogenesis; Background subtraction; Heterogeneity; Immuno-labeling; Mixture modeling; Platelet-Derived Growth Factor (PDGF); Quantitative flow cytometry; Systems biology; Vascular Endothelial Growth Factor (VEGF); qFlow cytometry