Elizabeth Stucky Hunter

Detecting Aspergillus -specific IgG is critical to diagnosing chronic pulmonary aspergillosis (CPA). Existing assays are often cost- and resource-intensive and not compatible with resource-constrained laboratory settings. LDBio Diagnostics has recently commercialized a lateral flow assay based on immunochromatographic technology (ICT) that detects Aspergillus antibodies (IgG and IgM) in less than 30 min, requiring minimal laboratory equipment.

Astroglia are well known for their role in propagating secondary injury following brain trauma. Modulation of this injury cascade, including inflammation, is essential to repair and recovery. Mesenchymal stromal cells (MSCs) have been demonstrated as trophic mediators in several models of secondary CNS injury, however, there has been varied success with the use of direct implantation due to a failure to persist at the injury site. To achieve sustained therapeutic benefit, we have encapsulated MSCs in alginate microspheres and evaluated the ability of these encapsulated MSCs to attenuate neuro-inflammation. In this study, astroglial cultures were administered lipopolysaccharide (LPS) to induce inflammation and immediately co-cultured with encapsulated or monolayer human MSCs. Cultures were assayed for the pro-inflammatory cytokine tumor necrosis factor alpha (TNF-[ ] produced by astroglia, MSC-produced prostaglandin E₂, and expression of neurotrophin-associated genes. We found that encapsulated MSCs significantly reduced TNF-[ ] produced by LPS-stimulated astrocytes more effectively than monolayer MSCs, and this enhanced benefit commences earlier than that of monolayer MSCs. Furthermore, in support of previous findings, encapsulated MSCs constitutively produced high levels of PGE₂, while monolayer MSCs required the presence of inflammatory stimuli to induce PGE₂production. The early, constitutive presence of PGE₂significantly reduced astrocyte-produced TNF-[ ], while delayed administration had no effect. Finally, MSC-produced PGE₂was not only capable of modulating inflammation, but appeared to have an additional role in stimulating astrocyte neurotrophin production. Overall, these results support the enhanced benefit of encapsulated MSC treatment, both in modulating the inflammatory response and providing neuroprotection.

Immunoassays are widely utilized due to their ability to quantify a vast assortment of biomolecules relevant to biological research and clinical diagnostics. Recently, immunoassay capabilities have been improved by the development of multiplex assays that simultaneously measure multiple analytes in a single sample. However, these assays are hindered by high costs of reagents and relatively large sample requirements. For example, in vitro screening systems currently dedicate individual wells to each time point of interest and this limitation is amplified in screening studies when the investigation of many experimental conditions is necessary; resulting in large volumes for analysis, a correspondingly high cost and a limited temporal experimental design. Microfluidics based immunoassays have been developed in order to overcome these drawbacks. Together, previous studies have demonstrated on-chip assays with either a large dynamic range, high performance sensitivity, and/or the ability to process samples in parallel on a single chip. In this report, we develop a multiplex immunoassay possessing all of these parallel characteristics using commercially available reagents, which allows the analytes of interest to be easily changed. The device presented can measure 6 proteins in 32 samples simultaneously using only 4.2 μL of sample volume. High quality standard curves are generated for all 6 analytes included in the analysis, and spiked samples are quantified throughout the working range of the assay. In addition, we demonstrate a strong correlation (R² = 0.8999) between in vitro supernatant measurements using our device and those obtained from a bench-top multiplex immunoassay. Finally, we describe cytokine secretion in an in vitro inflammatory hippocampus culture system, establishing proof-of-concept of the ability to use this platform as an in vitro screening tool. The low-volume, multiplexing abilities of the microdevice described in this report could be broadly applied to numerous situations where sample volumes and costs are limiting.