This review summarizes the significant biophysical and rheological aspects of red blood cell physiology and pathophysiology in relation to recent advances in microfluidic biomarker assays and ...emerging targeted or curative intent therapies.
Alterations in red cell biophysical properties and blood rheology have been associated with numerous hematologic and circulatory disorders. Recent advances in biomarker assays enable effective assessment of these biophysical and rheological properties in normoxia or physiological hypoxia in a clinically meaningful way. There are emerging targeted or curative therapies that aim to improve red cell pathophysiology, especially in the context of inherited hemoglobin disorders, such as sickle cell disease.
Red cell pathophysiology can be therapeutically targeted and the improvements in membrane and cellular biophysics and blood rheology can now be feasibly assessed via new microfluidic biomarker assays. Recent advances provide a new hope and novel treatment options for major red cell ailments, including inherited hemoglobin disorders, membrane disorders, and other pathologies of the red cell, such as malaria.
Optical sensor technology offers significant opportunities in the field of medical research and clinical diagnostics, particularly for the detection of small numbers of molecules in highly diluted ...solutions. Several methods have been developed for this purpose, including label-free plasmonic biosensors based on metamaterials. However, the detection of lower-molecular-weight (<500 Da) biomolecules in highly diluted solutions is still a challenging issue owing to their lower polarizability. In this context, we have developed a miniaturized plasmonic biosensor platform based on a hyperbolic metamaterial that can support highly confined bulk plasmon guided modes over a broad wavelength range from visible to near infrared. By exciting these modes using a grating-coupling technique, we achieved different extreme sensitivity modes with a maximum of 30,000 nm per refractive index unit (RIU) and a record figure of merit (FOM) of 590. We report the ability of the metamaterial platform to detect ultralow-molecular-weight (244 Da) biomolecules at picomolar concentrations using a standard affinity model streptavidin-biotin.
Sickle cell disease, a genetic disorder affecting a sizeable global demographic, manifests in sickle red blood cells (sRBCs) with altered shape and biomechanics. sRBCs show heightened adhesive ...interactions with inflamed endothelium, triggering painful vascular occlusion events. Numerous studies employ microfluidic-assay-based monitoring tools to quantify characteristics of adhered sRBCs from high resolution channel images. The current image analysis workflow relies on detailed morphological characterization and cell counting by a specially trained worker. This is time and labor intensive, and prone to user bias artifacts. Here we establish a morphology based classification scheme to identify two naturally arising sRBC subpopulations-deformable and non-deformable sRBCs-utilizing novel visual markers that link to underlying cell biomechanical properties and hold promise for clinically relevant insights. We then set up a standardized, reproducible, and fully automated image analysis workflow designed to carry out this classification. This relies on a two part deep neural network architecture that works in tandem for segmentation of channel images and classification of adhered cells into subtypes. Network training utilized an extensive data set of images generated by the SCD BioChip, a microfluidic assay which injects clinical whole blood samples into protein-functionalized microchannels, mimicking physiological conditions in the microvasculature. Here we carried out the assay with the sub-endothelial protein laminin. The machine learning approach segmented the resulting channel images with 99.1±0.3% mean IoU on the validation set across 5 k-folds, classified detected sRBCs with 96.0±0.3% mean accuracy on the validation set across 5 k-folds, and matched trained personnel in overall characterization of whole channel images with R2 = 0.992, 0.987 and 0.834 for total, deformable and non-deformable sRBC counts respectively. Average analysis time per channel image was also improved by two orders of magnitude (∼ 2 minutes vs ∼ 2-3 hours) over manual characterization. Finally, the network results show an order of magnitude less variance in counts on repeat trials than humans. This kind of standardization is a prerequisite for the viability of any diagnostic technology, making our system suitable for affordable and high throughput disease monitoring.
In recent years, considerable research efforts have been focused on near-perfect and perfect light absorption using metamaterials spanning frequency ranges from microwaves to visible frequencies. ...This relatively young field is currently facing many challenges that hampers its possible practical applications. In this paper, we present grating coupled-hyperbolic metamaterials (GC-HMM) as multiband perfect absorber that can offer extremely high flexibility in engineering the properties of electromagnetic absorption. The fabricated GC-HMMs exhibit several highly desirable features for technological applications such as polarization independence, wide angle range, broad- and narrow- band modes, multiband perfect and near perfect absorption in the visible to near-IR and mid-IR spectral range. In addition, we report a direct application of the presented system as an absorption based plasmonic sensor with a record figure of merit for this class of sensors.
This review briefly summarizes the significant impact of thromboinflammation in sickle cell disease in relation to recent advances in biomarkers that are used in functional microfluidic assays.
...Sickle cell disease (SCD) is an inherited hemoglobinopathy that affects 100 000 Americans and millions worldwide. Patients with SCD exhibit chronic haemolysis, chronic inflammation and thrombosis, and vaso-occlusion, triggering various clinical complications, including organ damage and increased mortality and morbidity. Recent advances in functional microfluidic assays provide direct biomarkers of disease, including abnormal white blood cell and red blood cell adhesion, cell aggregation, endothelial degradation and contraction, and thrombus formation.
Novel and emerging functional microfluidic assays are a promising and feasible strategy to comprehensively characterize thromboinflammatory reactions in SCD, which can be used for personalized risk assessment and tailored therapeutic decisions.
Seventy-five percent of patients with epithelial ovarian cancer present with advanced-stage disease that is extensively disseminated intraperitoneally and prognosticates the poorest outcomes. ...Primarily metastatic within the abdominal cavity, ovarian carcinomas initially spread to adjacent organs by direct extension and then disseminate via the transcoelomic route to distant sites. Natural fluidic streams of malignant ascites triggered by physiological factors, including gravity and negative subdiaphragmatic pressure, carry metastatic cells throughout the peritoneum. We investigated the role of fluidic forces as modulators of metastatic cancer biology in a customizable microfluidic platform using 3D ovarian cancer nodules. Changes in the morphological, genetic, and protein profiles of biomarkers associated with aggressive disease were evaluated in the 3D cultures grown under controlled and continuous laminar flow. A modulation of biomarker expression and tumor morphology consistent with increased epithelial–mesenchymal transition, a critical step in metastatic progression and an indicator of aggressive disease, is observed because of hydrodynamic forces. The increase in epithelial–mesenchymal transition is driven in part by a posttranslational up-regulation of epidermal growth factor receptor (EGFR) expression and activation, which is associated with the worst prognosis in ovarian cancer. A flow-induced, transcriptionally regulated decrease in E-cadherin protein expression and a simultaneous increase in vimentin is observed, indicating increased metastatic potential. These findings demonstrate that fluidic streams induce a motile and aggressive tumor phenotype. The microfluidic platform developed here potentially provides a flow-informed framework complementary to conventional mechanism-based therapeutic strategies, with broad applicability to other lethal malignancies.
We present a microfluidic approach that allows simultaneous interrogation of RBC properties in physiological flow conditions at a single cell level. With this method, we studied healthy hemoglobin A ...(HbA) and homozygous sickle hemoglobin (HbS) containing RBCs using whole blood samples from twelve subjects. We report that HbS-containing RBCs are heterogeneous in terms of adhesion and deformability in flow.
Objective: Effective management of diabetes largely benefits from early diagnosis followed by intensive long-term regulation of blood glucose. The levels of glycohemoglobin (HbA1 and HbA1c) have been ...used as standard biomarkers to assess long-term blood glucose concentrations for diabetes diagnosis and management. Gold standard laboratory methods for HbA1 and HbA1c testing are often costly and not widely available. Moreover, currently available point-of-care (POC) immunoassay-based glycohemoglobin tests may produce inaccurate test results for patients with co-existing diseases such as hemoglobin disorders and anemia. Here, we report a POC platform, HemeChip-GHb, for quantitative HbA1 detection leveraging paper-based affinity electrophoresis. Methods: We describe the design and development of the HemeChip-GHb test. Feasibility and accuracy of the HemeChip-GHb system were demonstrated by testing blood samples collected from healthy donors, patients with prediabetes, and patients with diabetes. Results: HbA1 levels measured with HemeChip-GHb show 0.96 correlation to the levels reported from the clinical standard HPLC tests, and with a bias of -0.72% based on Bland-Altman analysis. 99.6% of the HbA1 levels for paired HemeChip-GHb and HPLC fell within A and B zones of no difference in clinical outcome based on error grid analysis. Conclusion: Using HemeChip-GHb we achieved accurate diabetes status detection with sensitivity and specificity of 100%. Significance: We presented a novel POC paper-based affinity electrophoresis platform that has the potential for accurately diagnosing diabetes, and addressing an unmet need for accurate and affordable diagnostics in resource-challenged environments.
Alterations in the deformability of red blood cells (RBCs), occurring in hemolytic blood disorders such as sickle cell disease (SCD), contribute to vaso-occlusion and disease pathophysiology. There ...are few functional in vitro assays for standardized assessment of RBC-mediated microvascular occlusion. Here, we present the design, fabrication, and clinical testing of the Microfluidic Impedance Red Cell Assay (MIRCA) with embedded capillary network-based micropillar arrays and integrated electrical impedance measurement electrodes to address this need. The micropillar arrays consist of microcapillaries ranging from 12 μm to 3 μm, with each array paired with two sputtered gold electrodes to measure the impedance change of the array before and after sample perfusion through the microfluidic device. We define RBC occlusion index (ROI) and RBC electrical impedance index (REI), which represent the cumulative percentage occlusion and cumulative percentage impedance change, respectively. We demonstrate the promise of MIRCA in two common red cell disorders, SCD and hereditary spherocytosis. We show that the electrical impedance measurement reflects the microvascular occlusion, where REI significantly correlates with ROI that is obtained via high-resolution microscopy imaging of the microcapillary arrays. Further, we show that RBC-mediated microvascular occlusion, represented by ROI and REI, associates with clinical treatment outcomes and correlates with in vivo hemolytic biomarkers, lactate dehydrogenase (LDH) level and absolute reticulocyte count (ARC) in SCD. Impedance measurement obviates the need for high-resolution imaging, enabling future translation of this technology for widespread access, portable and point-of-care use. Our findings suggest that the presented microfluidic design and the integrated electrical impedance measurement provide a reproducible functional test for standardized assessment of RBC-mediated microvascular occlusion. MIRCA and the newly defined REI may serve as an in vitro therapeutic efficacy benchmark for assessing the clinical outcome of emerging RBC-modifying targeted and curative therapies.
Commercial off-the-shelf digital cameras are inexpensive and easy-to-use instruments that can be used for quantitative scientific data acquisition if images are captured in raw format and processed ...so that they maintain a linear relationship with scene radiance. Here we describe the image-processing steps required for consistent data acquisition with color cameras. In addition, we present a method for scene-specific color calibration that increases the accuracy of color capture when a scene contains colors that are not well represented in the gamut of a standard color-calibration target. We demonstrate applications of the proposed methodology in the fields of biomedical engineering, artwork photography, perception science, marine biology, and underwater imaging.
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