In VitroNeutrophil-Bacteria Assay in Whole Blood Microenvironments with Single-Cell Confinement

Abstract

Blood is a common medium through which invasive bacterial infections disseminate in the human body. In vitro neutrophil-bacteria assays allow flexible mechanistic studies and screening of interventional strategies. In standard neutrophil-bacteria assays, both the immune cells and microorganisms are typically interrogated in an exogenous, homogeneous, bulk fluid environment (e.g., culture media or bacterial broth in microtiter plates), lacking the relevant physicochemical factors in the heterogenous blood-tissue microenvironment (e.g., capillary bed) with single-cell confinement. Here we present an in vitro neutrophil-bacteria assay by leveraging an open microfluidic model known as μ-Blood that supports sub-microliter liquid microchannels with single-cell confinement. In this study we compare the exogenous and endogenous fluids including neutrophils in RPMI (standard suspension cell culture media) and whole blood in response to Staphylococcus aureus (S. aureus, a gram-positive, non-motile bacterium) in phosphate buffered saline (PBS), Mueller Hinton Broth (MHB), and human serum. Our results reveal a significant disparity between the exogenous and endogenous fluid microenvironments in the growth kinetics of bacteria, the spontaneous generation of capillary (i.e., Marangoni) flow, and the outcome of neutrophil intervention on the spreading bacteria.

Combining Top-Down and Bottom-Up: An Open Microfluidic Microtumor Model for Investigating Tumor Cell-ECM Interaction and Anti-Metastasis

“…This UOMS microtumor model can be further integrated with other UOMS-based technologies including autonomously regulated oxygen microenvironment (AROM), [12] microliter whole blood assay (μ-Blood), [44,45] microbial communities, [26,46] ELR-antimicrobial susceptibility testing (AST), [27] micro total analysis system (μTAS), [22] and label-free spectroscopies (e.g., multiphoton, [26] Raman, [28] and IR [47] ). Thanks to its natural alignment with the open system standard, the UOMS microtumor model is compatible with most the automated liquid handling tools (e.g., robotic pipetting/microtiter plate preparation systems) in biology and clinical laboratories.…”