Simplified size-based immune cell monitoring using a hand-held particle counting device

Abstract

Abstract Biological samples are often heterogeneous cell mixtures that differ by type and functional or responsive capacity. Beyond differences in protein expression, many cell types and physiological states are also distinguishable by size (cell diameter). The ability to distinguish population subsets is critical to many aspects of research. Such variation in cellular attributes is most commonly determined by multicolor fluorescent antibody detection of cell type-specific surface markers using flow cytometry. While flow cytometers are complex, trained operator-dependent platforms of core facilities, the availability of simplified methods for identifying activation status and measuring differences in the relative cell frequencies of more than one cell type in samples or co-cultures is somewhat limited. Moreover, despite the need for speed and accuracy, hemocytometry, the most widely used technique for cell counting, is quite tedious and fraught with user-driven variability. To address these needs, we present application of a hand-held cell counter that operates on the basic Coulter Principle of measuring changes in electrical impedance produced by particles (cells or beads) as they pass through a defined aperture. Specifically, instrumentation has been collapsed to the size of a pipette; the precision-made, microfluidic sensor contains the cell-sensing zone that enables size discrimination at the submicron with high counting accuracy. To demonstrate performance, the platform was used to assess cell distribution in human PBMC isolates and determine immune cell activation in response to CD3/CD28 co-stimulation or PHA exposure. All experiments were conducted in comparison to flow cytometry and a standard Coulter device.