Neutropenia is a condition identified by an abnormally low number of neutrophils in the bloodstream and signifies an increased risk of severe infection. Cancer patients are particularly susceptible to this condition, which can be disruptive to their treatment and even life-threatening in severe cases. Thus, it is critical to routinely monitor neutrophil counts in cancer patients. However, the standard of care to assess neutropenia, the complete blood count (CBC), requires expensive and complex equipment, as well as cumbersome procedures, which precludes easy or timely access to critical hematological information, namely neutrophil counts. Here we present a simple, low-cost, fast, and robust technique to detect and grade neutropenia based on label-free multi-spectral deep-UV microscopy. Results show that the developed framework for automated segmentation and classification of live, unstained blood cells in a smear accurately differentiates patients with moderate and severe neutropenia from healthy samples in minutes. This work has significant implications towards the development of a low-cost and easy-to-use point-of-care device for tracking neutrophil counts, which can not only improve the quality of life and treatment-outcomes of many patients but can also be lifesaving.

The correlation between cardiovascular disease and iron deficiency anemia (IDA) is well documented but poorly understood. Using a multi-disciplinary approach, we explore the hypothesis that the biophysical alterations of red blood cells (RBCs) in IDA, such as variable degrees of microcytosis and decreased deformability may directly induce endothelial dysfunction via mechanobiological mechanisms. Using a combination of atomic force microscopy and microfluidics, we observed that subpopulations of IDA RBCs (idRBCs) are significantly stiffer and smaller than both healthy RBCs and the remaining idRBC population. Furthermore, computational simulations demonstrated that the smaller and stiffer idRBC subpopulations marginate toward the vessel wall causing aberrant shear stresses. This leads to increased vascular inflammation as confirmed with perfusion of idRBCs into our “endothelialized” microfluidic systems. Overall, our multifaceted approach demonstrates that the altered biophysical properties of idRBCs directly lead to vasculopathy, suggesting that the IDA and cardiovascular disease association extends beyond correlation and into causation.

Hematological analysis, via a complete blood count (CBC) and microscopy, is critical for screening, diagnosing, and monitoring blood conditions and diseases but requires complex equipment, multiple chemical reagents, laborious system calibration and procedures, and highly trained personnel for operation. Here we introduce a hematological assay based on label-free molecular imaging with deep-ultraviolet microscopy that can provide fast quantitative information of key hematological parameters to facilitate and improve hematological analysis. We demonstrate that this label-free approach yields 1) a quantitative five-part white blood cell differential, 2) quantitative red blood cell and hemoglobin characterization, 3) clear identification of platelets, and 4) detailed subcellular morphology. Analysis of tens of thousands of live cells is achieved in minutes without any sample preparation. Finally, we introduce a pseudocolorization scheme that accurately recapitulates the appearance of cells under conventional staining protocols for microscopic analysis of blood smears and bone marrow aspirates. Diagnostic efficacy is evaluated by a panel of hematologists performing a blind analysis of blood smears from healthy donors and thrombocytopenic and sickle cell disease patients. This work has significant implications toward simplifying and improving CBC and blood smear analysis, which is currently performed manually via bright-field microscopy, and toward the development of a low-cost, easy-to-use, and fast hematological analyzer as a point-of-care device and for low-resource settings.

Investigating individual red blood cells (RBCs) is critical to understanding hematologic diseases, as pathology often originates at the single-cell level. Many RBC disorders manifest in altered biophysical properties, such as deformability of RBCs. Due to limitations in current biophysical assays, there exists a need for high-throughput analysis of RBC deformability with single-cell resolution. To that end, we present a method that pairs a simple in vitro artificial microvasculature network system with an innovative MATLAB-based automated particle tracking program, allowing for high-throughput, single-cell deformability index (sDI) measurements of entire RBC populations. We apply our technology to quantify the sDI of RBCs from healthy volunteers, Sickle cell disease (SCD) patients, a transfusion-dependent beta thalassemia major patient, and in stored packed RBCs (pRBCs) that undergo storage lesion over 4 weeks. Moreover, our system can also measure cell size for each RBC, thereby enabling 2D analysis of cell deformability vs cell size with single cell resolution akin to flow cytometry. Our results demonstrate the clear existence of distinct biophysical RBC subpopulations with high interpatient variability in SCD as indicated by large magnitude skewness and kurtosis values of distribution, the “shifting” of sDI vs RBC size curves over transfusion cycles in beta thalassemia, and the appearance of low sDI RBC subpopulations within 4 days of pRBC storage. Overall, our system offers an inexpensive, convenient, and high-throughput method to gauge single RBC deformability and size for any RBC population and has the potential to aid in disease monitoring and transfusion guidelines for various RBC disorders.