There has been increased interest in using mechanical properties of cells as label-free biomarkers for applications in basic biology and clinical diagnostics. Increased single-cell deformability has been correlated with malignancy in cell lines and patient samples for example. In contrast to conventional molecular biomarkers a mechanical property is intrinsic to the cells of interest and does not require extensive sample preparation or costly labels. Still, clinical applications in this field have been limited to the research setting as previous tools have lacked the throughput to examine the heterogeneity of complex clinical samples effectively. The proposed work builds upon the deformability cytometry platform, which employs inertial microfluidic principles to create a fluid shearing extensional flo junction that can analyze mechanical properties of over 1000 cells per second. At the research platform stage, the deformability cytometry system has shown very promising clinical diagnostic utility in identifying malignant cells in pleural fluid with much higher sensitivity than the gold standard cytological analysis. Here we build off this initial success and address two major challenges to create a commercially viable high- throughput mechanical phenotyping platform. We aim to reduce costs associated with high-speed camera readout, and decrease computational resources required to achieve near-real-time-rapid-decision analysis, faster data turnaround time and downstream sorting capabilities. Specifically, the aims of this proposal are to explore the feasibility of lower cost single-point interrogation modalities and high-speed imaging alternatives while retaining similar diagnostic performance. Single-point interrogation methods include transit-time of a pre- deformed cell prior to mechanically applied stress compared to post-deformed cells. Alternatively, single-point measurements of scattered light during the deformation events, similar to side-scatter flow cytometry measurements will be correlated to deformed cell shape. Both these approaches are expected to reduce computational resources as output data is 1D. Three alternative high-speed imaging techniques will also be evaluated 1) Position sensitive detectors (PSDs) are one axis imaging sensors - by placing two PSDs orthogonally 2D spatial data can be collected during deformation events, 2) Commercially available OEM high- speed image sensors. are customizable where onboard FPGA integration can be used to prescreen data in order to reduce offline image analysis workload, 3) adapted lower frame rate camera (10,000 fps) coupled to 500ns strobe source. Any of the proposed alternative acquisition methods will reduce the cost of the instrument and lower analysis time to create a clinically relevant label-free mechanical biomarker-based diagnostic product.
Public Health Relevance Statement: Public Health Relevance: We are developing a next generation diagnostic tool to identify cancer in body fluids based on measuring the inherent mechanical features of the cells. The proposed work will characterize reduced complexity methods of recording cell mechanical properties for a commercial diagnostic instrument. Ultimately, this tool will improve diagnostic accuracy with a simple automated platform, minimize the number of invasive procedures, and reduce overall healthcare costs.
Project Terms: Acute; Address; analog; Area; base; Benign; Biological Assay; Biological Markers; Biology; Biopsy; Body Fluids; cancer cell; care burden; Cell Line; Cell Shape; cell type; Cells; Characteristics; Chest; Chronic; Clinical; clinical application; Clinical Research; clinically relevant; Complement; Complex; Computer Analysis; computing resources; cost; cost effective; Coupled; Cytology; Cytometry; Data; data acquisition; Data Set; Decision Analysis; detector; Diagnosis; Diagnostic; diagnostic accuracy; Diagnostic Sensitivity; Disease; Engineering; Event; Flow Cytometry; Future; Goals; Gold; granulocyte; Health Care Costs; Healthcare; Heterogeneity; high risk; Image; Image Analysis; Imaging Techniques; Immune; Immunohistochemistry; improved; In Vitro; Individual; Inflammation; Inflammatory; instrument; interest; Label; Lead; Leukocytes; Liquid substance; Low Prevalence; Lung; Malignant - descriptor; Malignant Neoplasms; Malignant Pleural Effusion; Masks; Measurement; Measures; Mechanics; Medicine; Mesothelial Cell; Methods; Microfluidics; Modality; Molecular; Monitor; Neoplasm Metastasis; nephelometry; neutrophil; novel diagnostics; Outcome; Output; Patients; Performance; Phase; Phenotype; Pleural; Pleural cavity; Pleural effusion disorder; Population; Population Heterogeneity; Positioning Attribute; Preparation; pressure; Procedures; Process; Property; prototype; public health relevance; Recording of previous events; Research; Risk; Sampling; screening; sensor; Side; Sorting - Cell Movement; Source; Specificity; Speed (motion); Staging; Stress; success; System; Techniques; Thoracentesis; Time; tool; Work; Workload
