Sample-to-Answer Immuno-Magnetic Assay Using Thermally Responsive Alkane Partitions.

To combat pandemics, there is a need for rapid point-of-care diagnostics to identify infected patients and to track the spread of the disease. While recent progress has been made in response to COVID-19, there continues to be a need for point-of-care diagnostics capable of detecting biomarkers-such as antibodies-in whole blood. We have recently reported the development of thermally responsive alkane partitions (TRAPs) for the automation of point-of-care immuno-magnetic assays. Here, we demonstrate the use of TRAPs to enable sample-to-answer detection of antibodies against the SARS-CoV-2 virus in whole blood samples. We report a limit of detection of 84 pg/mL, well below the clinically relevant threshold. We anticipate that the TRAP-enabled sample-to-answer immunoassay can be used to track the progression of future pandemics, leading to a more informed and robust clinical and societal response.

Thermally Responsive Alkane Partitions for Assay Automation.

For point-of-care diagnostic tools to be impactful, they must be inexpensive, equipment-free, and sample-to-answer (i.e., require no user intervention). Here, we report a new approach to enable sample-to-answer diagnostics that utilizes thermally responsive alkane partitions (TRAPs) as automated pseudo-valves. When combined with the magnetic manipulation of microbeads, TRAPs enable the pumpless automation of all steps in complex assays. We demonstrate that in relatively narrow channel geometries, liquified alkane partitions continue to separate reagents on each side of the partition while enabling the transition of magnetic beads from one reagent to the next, replacing manual pipetting steps in conventional assays. In addition, we show that in relatively broader geometries, liquified partitions breach, enabling the addition/mixing of preloaded reagents. Through calculation and experimentation, we determine the geometric design rules for implementing the stationary and removable partitions in fluidic channels. In addition, we demonstrate that magnetic microbeads can be pulled through liquified stationary TRAPs without disrupting partition integrity and without disrupting bound protein complexes attached at the microbead surface. The TRAP technology introduced here can enable a new low-cost and equipment-free approach for fully automated sample-to-answer diagnostics.

Sample-to-Answer Diagnostic System for the Detection of Circulating Histones in Whole Blood.

Severe internal trauma results in millions of hospitalizations each year, including thousands of deaths caused by subsequent multiple organ failure. The majority of these deaths occur within the first 24 h, and thus, rapid diagnosis of internal trauma severity is necessary for immediate treatment. For early organ damage identification, diagnosis in point-of-care settings is crucial for rapid triage and treatment. Recent reports suggest that circulating histones may serve as a biomarker for severe organ damage and the risk of multiple organ failure. Here, we report a point-of-care diagnostic system that utilizes the inherent interactions between histones and DNA for the fluorescence-based detection of histones in whole blood. In the assay, histones within the sample are wrapped by DNA, thus preventing an intercalating dye from binding the DNA and fluorescing. To allow for quantitative fluorescent measurements to be made in a point-of-care setting, we integrate a rapid, automated blood separation step into our assay. Furthermore, we eliminate manual reagent additions using a thermally responsive alkane partition (TRAP), thus making the system sample-to-answer. Finally, we demonstrate the assay in a portable fluorescence reader compatible with a point-of-care environment. We report a limit of detection 112 ng/mL in whole blood, suggesting that our device can be used to rapidly diagnose internal trauma severity and the likelihood of multiple organ failure in near-patient settings.